Compositions and methods for plasmapheresis

ABSTRACT

Described herein are compositions and methods for performing plasmapheresis. The compositions and methods for performing plasmapheresis are innovative at least in their application towards the treatment and prevention of aging and conditions associated with aging. Plasmapheresis compositions and methods described herein are directed towards reducing or eliminating conditions associated with aging.

CROSS-REFERENCE

This application claims the benefit of U.S. Provisional PatentApplication No. 63/347,124, which was filed on May 31, 2022, and whichis incorporated herein in its entirety by reference.

BACKGROUND OF THE DISCLOSURE

During the past century, the earth's population has more than doubled.It is estimated that more than 20% of the world's population is aged 65years or older. The United Nations estimates that, by 2050, thispopulation will have increased beyond 14 billion. Aged humans almostinevitably suffer from one or more disorders associated with chronicaging. These can include Alzheimer's disease, infections, Type IIDiabetes, atherosclerotic cardio vascular disease, obesity,osteoporosis, and sarcopenia. The cumulative effect is an enormousfinancial burden to any medical system.

SUMMARY OF THE DISCLOSURE

Described herein is a method of providing plasmapheresis to anindividual in order to improve a health status of the individual,comprising steps of: (a) measuring, before an administration ofplasmapheresis, one or more of a strength of the individual, a balanceof the individual, a mental status of the individual, and a measure of awalking of the individual, thereby generating an indication of apre-treatment health status of the individual; (b) administering theplasmapheresis to the individual; (c) measuring, following step (b), oneor more of the strength of the individual, the balance of theindividual, the mental status of the individual, and the measure of thewalking of the individual, thereby generating an indication of apost-treatment health status of the individual; and (d) comparing thepost-treatment health status of the individual with the pre-treatmenthealth status of the individual in order to determine that the healthstatus of the individual has improved as a result of the plasmapheresisadministration. In some embodiments, step (a) occurs within 24 hours ofthe administering of the plasmapheresis to the individual in step (b).In some embodiments, the strength of the individual is measured in step(b) by measuring a grip strength of the individual. In some embodiments,balance of the individual is measured in step (a), step (c), or steps(a) and (c) by having the individual stand on one leg and measuring howlong the individual remains standing with one leg raised. In someembodiments, the mental status is measured in step (a), step (c), orsteps (a) and (c) using a survey comprising questions that assessemotional wellbeing. In some embodiments, the measure of the walking ofthe individual is measured in step (a), step (c), or steps (a) and (c)by having the individual stand from a seated position and walk. In someembodiments, the plasmapheresis that is administered in step (b)exchanges at least one unit of plasma volume. In some embodiments, theplasmapheresis that is administered in step (b) is administered over aplurality of treatment sessions. In some embodiments, two treatmentsessions of the plurality of treatment sessions are administered within72 hours of each other. In some embodiments, the plasmapheresis that isadministered in step (b) is administered over a single treatmentsession. In some embodiments, the comparing the post-treatment healthstatus of the individual with the pre-treatment health status of theindividual in step (d) results in a determination of a quantitativedifference between the post-treatment health status of the individualand the pre-treatment health status of the individual. In someembodiments, the method further comprises repeating steps (b)-(d) untilthe quantitative difference is a specific value. In some embodiments,the method further comprises repeating steps (a)-(d) until thequantitative difference is a specific value.

Described herein is a method for treating a condition associated withaging in an individual, comprising: (a) administering plasmapheresis tothe individual; and (b) monitoring for a change in the condition. Insome embodiments, the condition associated with aging comprises a lossof strength. In some embodiments, the strength of the individual ismonitored in step (b) by measuring a grip strength of the individual. Insome embodiments, the condition associated with aging comprises a lossof balance. In some embodiments, balance of the individual is monitoredin step (b) by having the individual stand on one leg and measuring howlong the individual remains standing with one leg raised. In someembodiments, the condition associated with aging comprises diminishedability to walk. In some embodiments, the ability to walk of theindividual is monitored in step (b) by having the individual stand froma seated position and walk. In some embodiments, the plasmapheresis thatis administered in step (a) exchanges at least one unit of plasmavolume. In some embodiments, the plasmapheresis that is administered instep (a) is administered over a plurality of treatment sessions. In someembodiments, two treatment sessions of the plurality of treatmentsessions are administered within 72 hours of each other. In someembodiments, the plasmapheresis that is administered in step (a) isadministered over a single treatment session. In some embodiments, themethod further comprises repeating steps (a)-(b) until the change in thecondition is achieved.

Described is a method for using plasmapheresis to treat an individual byusing the plasmapheresis to modulate an amount of expression of a cellsurface marker on the cell surface of a white blood cell of theindividual, comprising the steps of: (a) measuring, before anadministration of plasmapheresis, a level of expression of the cellsurface marker in blood of the individual; (b) administering theplasmapheresis to the individual; and (c) measuring, following step (b),the level of expression of the cell surface marker in the blood of theindividual and determining that the expression of the cell surfacemarker on the cell surface of the white blood cell of the individual hasbeen modulated. In some embodiments, the white blood cell comprises alymphocyte. In some embodiments, the lymphocyte comprises a T-cell. Insome embodiments, the white blood cell comprises a monocyte. In someembodiments, the white blood cell comprises a basophil. In someembodiments, the white blood cell comprises a neutrophil. In someembodiments, the white blood cell comprises an eosinophil. In someembodiments, to modulate the amount of expression of the cell surfacemarker on the cell surface of the white blood cell of the individual isto change the amount of expression of the cell surface marker to adegree that is measurable in the blood of the individual following theadministering of plasmapheresis in step (b). In some embodiments, thecell surface marker comprises CD16. In some embodiments, the cellsurface marker comprises CD25. In some embodiments, the cell surfacemarker comprises CD27. In some embodiments, the cell surface markercomprises CD38. In some embodiments, the cell surface marker comprisesCD57. In some embodiments, the cell surface marker comprises CD80. Insome embodiments, the cell surface marker comprises HLADR. In someembodiments, the cell surface marker comprises IgM. In some embodiments,the cell surface marker comprises KTR. In some embodiments, the cellsurface marker comprises KLRG1. In some embodiments, the cell surfacemarker comprises NK1. In some embodiments, the cell surface markercomprises NKg2a. In some embodiments, the cell surface marker comprisesTIGIT. In some embodiments, step (a) occurs within 24 hours of theadministering of the plasmapheresis to the individual in step (b). Insome embodiments, the plasmapheresis that is administered in step (b)exchanges at least one unit of plasma volume. In some embodiments, theplasmapheresis that is administered in step (b) is administered over aplurality of treatment sessions. In some embodiments, two treatmentsessions of the plurality of treatment sessions are administered within72 hours of each other. In some embodiments, the plasmapheresis that isadministered in step (b) is administered over a single treatmentsession. In some embodiments, the expression of the cell surface markeris measured using flow cytometry. In some embodiments, the expression ofthe cell surface marker is measured using a fluorescent conjugatedantibody. In some embodiments, the modulation is a measurable decreasebetween level of expression of the cell surface marker that is measuredin step (a) and the level of expression of the cell surface markermeasured in step (c). In some embodiments, the method further comprisesrepeating steps (b)-(c) until the cell surface modulation having aspecific value is achieved. In some embodiments, the method furthercomprising repeating steps (a)-(c) until the cell surface modulationhaving a specific value is achieved.

Described herein is a method for treating aging in an individual byusing plasmapheresis to reduce cellular senescence in the individual,comprising the steps of: (a) measuring, before an administration ofplasmapheresis, a level of expression of a marker associated with thecellular senescence in blood of the individual; (b) administering theplasmapheresis to the individual; and (c) measuring, following step (b),the level of expression of the marker associated with the cellularsenescence and determining that the cellular senescence in theindividual has been reduced. In some embodiments, a cell in which thecellular senescence is reduced comprises a lymphocyte. In someembodiments, the lymphocyte comprises a T-cell. In some embodiments, acell in which the cellular senescence is reduced comprises a monocyte.In some embodiments, a cell in which the cellular senescence is reducedcomprises a basophil. In some embodiments, a cell in which the cellularsenescence is reduced comprises a neutrophil. In some embodiments, acell in which the cellular senescence is reduced comprises aneosinophil. In some embodiments, the marker associated with the cellularsenescence comprises senescence-associated beta-galactosidase(“SA-β-gal”). In some embodiments, step (a) occurs within 24 hours ofthe administering of the plasmapheresis to the individual in step (b).In some embodiments, the plasmapheresis that is administered in step (b)exchanges at least one unit of plasma volume. In some embodiments, theplasmapheresis that is administered in step (b) is administered over aplurality of treatment sessions. In some embodiments, two treatmentsessions of the plurality of treatment sessions are administered within72 hours of each other. In some embodiments, the plasmapheresis that isadministered in step (b) is administered over a single treatmentsession. In some embodiments, the expression of the marker associatedwith the cellular senescence is measured using flow cytometry. In someembodiments, the expression of the marker associated with the cellularsenescence is measured using a fluorescent conjugated antibody. In someembodiments, the modulation is a measurable decrease between level ofexpression of the marker associated with the cellular senescence that ismeasured in step (a) and the level of expression of the markerassociated with the cellular senescence that is measured in step (c). Insome embodiments, the method further comprises repeating steps (b)-(c)until a specific value is achieved for the reduction of the level ofexpression of the marker associated with the cellular senescence. Insome embodiments, the method further comprises repeating steps (a)-(c)until a specific value is achieved for the reduction of the level ofexpression of the marker associated with the cellular senescence.

Described herein is a method for performing plasmapheresis for use intreating or preventing a condition that is associated with aging in anindividual, the method comprising: removing, from within a vascularsystem of an individual, at least 70% of a factor that is associatedwith aging by performing plasmapheresis on the individual at least twotimes within a 72 hour period and thereby treating or preventing thecondition that is associated with aging in the individual. In someembodiments of the method, each of the at least two times thatplasmapheresis is performed comprises removing at least one plasmavolume from the individual. In some embodiments of the method, a volumeof exchange fluid that is returned to the individual is equal in volumeto the at least one plasma volume that is withdrawn. In some embodimentsof the method, a volume of exchange fluid that is returned to theindividual is greater in volume than the at least one plasma volume thatis withdrawn. In some embodiments of the method, at least one of the atleast two times that plasmapheresis is performed comprises removing atleast one- and one-half plasma volumes from the individual. In someembodiments of the method, a volume of exchange fluid that is returnedto the individual is equal to the at least one- and one-half plasmavolumes that is withdrawn. In some embodiments of the method, a volumeof exchange fluid that is returned to the individual is greater than theat least one- and one-half plasma volume that is withdrawn. In someembodiments of the method, the plasmapheresis includes infusing anexchange fluid into a vascular system of the individual and wherein theexchange fluid comprises at least one of: saline, Lactated Ringer's,albumin, or therapeutic. In some embodiments of the method, thetherapeutic comprises at least one of: an anti-inflammatory or animmune-modulator. In some embodiments of the method, theimmune-modulator comprises intravenous immunoglobulin. In someembodiments of the method, the method comprises the step ofadministering a therapeutic to the individual following at least one ofthe at least two times that the plasmapheresis is performed. In someembodiments of the method, the therapeutic comprises at least one of: ananti-inflammatory or an immune-modulator. In some embodiments of themethod, the immune-modulator comprises intravenous immunoglobulin.

Described herein is a method for performing plasmapheresis for use intreating or preventing a condition that is associated with aging in anindividual, the method comprising: (a) determining a biological age ofan individual; (b) performing plasmapheresis on the individual; and (c)repeating steps (a) and (b) until the biological age of the individualis below a threshold value. In some embodiments of the method, thebiological age of the individual is determined using an albumin bloodlevel of the individual. In some embodiments of the method, thebiological age of the individual is determined using a degree ofglycation of albumin in blood of the individual. In some embodiments ofthe method, the biological age of the individual is determined using aceruloplasmin blood level of the individual. In some embodiments of themethod, the biological age of the individual is determined using a levelof an immunoglobulin in the blood of the individual. In some embodimentsof the method, the biological age of the individual is determined usinga glutathione blood level of the individual. In some embodiments of themethod, the biological age of the individual is determined using anantibody assay, and wherein the antibody assay comprises at least one ofan antinuclear antibody screen, a rheumatoid factor assay, a thyroidperoxidase antibody assay, or a quantitative immunoglobulin assay. Insome embodiments of the method, the biological age of the individual isdetermined using a proteomic assay, and wherein the proteomic assaycomprises at least one of a fibrinogen assay, a creatinine kinase assay,or a hemoglobin A1C assay. In some embodiments of the method, thebiological age of the individual is determined using a metabolomicassay, and wherein the metabolomic assay comprises at least one of acholesterol assay or a blood glucose assay. In some embodiments of themethod, the biological age of the individual is determined using aurinalysis. In some embodiments of the method, the biological age of theindividual is determined using a peripheral blood mononuclear cellanalysis. In some embodiments of the method, the biological age of theindividual is determined using a cellular senescence assay. In someembodiments of the method, the biological age of the individual isdetermined using a genomic methylation assay. In some embodiments of themethod, the biological age of the individual is determined using aninflammatory marker analysis. In some embodiments of the method, thebiological age of the individual is determined using at least one of acomplete blood count, a total protein assay, a liver function assay, ablood urea nitrogen assay, a creatinine assay, or a c-reactive proteinassay.

Described herein is a method for performing a plasmapheresis regimen,comprising the steps of: (a) withdrawing at least 1 plasma volume ofwhole blood from an individual; (b) separating the whole blood into acellular fraction and a plasma fraction; (c) returning the cellularfraction to the individual; (d) infusing an exchange fluid to theindividual simultaneously with step (a); and (e) repeating steps (a)-(d)until at least one component found in plasma of the individual isdiluted by at least 60% as compared to before the plasmapheresis regimenwas initiated.

Described herein is a method for treating a condition associated withaging in an individual, comprising performing plasmapheresis on theindividual and removing at least one plasma volume from the individualduring the plasmapheresis. In some embodiments, the condition associatedwith aging is a decrease in strength of the individual. In someembodiments, wherein the condition associated with aging is a decreasein ambulation of the individual. In some embodiments, the conditionassociated with aging is a decrease in balance in the individual. Insome embodiments, the condition associated with aging is a decrease ofthe mental status of the individual. In some embodiments, the conditionassociated with aging is an increase inflammation in the individual. Insome embodiments, the increase in the inflammation in the individual isassociated with a change in level of expression of a cell surfaceprotein expressed on the surface of a white blood cell. In someembodiments, the white blood cell comprises a lymphocyte. In someembodiments, the lymphocyte comprises a T-cell. In some embodiments, thewhite blood cell comprises a monocyte. In some embodiments, the whiteblood cell comprises a basophil. In some embodiments, the white bloodcell comprises a neutrophil. In some embodiments, the white blood cellcomprises an eosinophil. In some embodiments, the cell surface proteincomprises CD16. In some embodiments, the cell surface protein comprisesCD25. In some embodiments, the cell surface protein comprises CD27. Insome embodiments, the cell surface protein comprises CD38. In someembodiments, the cell surface protein comprises CD57. In someembodiments, the cell surface protein comprises CD80. In someembodiments, the cell surface protein comprises HLADR. In someembodiments, the cell surface protein comprises IgM. In someembodiments, the cell surface protein comprises KTR. In someembodiments, the cell surface protein comprises KLRG1. In someembodiments, the cell surface protein comprises NK1. In someembodiments, the cell surface marker comprises NKg2a. In someembodiments, the cell surface protein comprises TIGIT.

A plasmapheresis exchange fluid composition for use in administeringplasmapheresis for treating or preventing a condition that is associatedwith aging in an individual, wherein a total volume of theplasmapheresis exchange fluid composition is equal to at least oneplasma volume of the individual. In some embodiments, the compositioncomprises 5% albumin. In some embodiments, the composition comprisesIVIG. In some embodiments, the condition associated with aging is adecrease in strength of the individual. In some embodiments, thecondition associated with aging is a decrease in ambulation of theindividual. In some embodiments, the condition associated with aging isa decrease in balance in the individual. In some embodiments, thecondition associated with aging is a decrease of the mental status ofthe individual. In some embodiments, the condition associated with agingis an increase inflammation in the individual. In some embodiments, theincrease in the inflammation in the individual is associated with achange in a level of expression of a cell surface protein expressed onthe surface of a white blood cell. In some embodiments, the white bloodcell comprises a lymphocyte. In some embodiments, the lymphocytecomprises a T-cell. In some embodiments, the white blood cell comprisesa monocyte. In some embodiments, the white blood cell comprises abasophil. In some embodiments, the white blood cell comprises aneutrophil. In some embodiments, the white blood cell comprises aneosinophil. In some embodiments, the cell surface protein comprisesCD16. In some embodiments, the cell surface protein comprises CD25. Insome embodiments, the cell surface protein comprises CD27. In someembodiments, the cell surface protein comprises CD38. In someembodiments, the cell surface protein comprises CD57. In someembodiments, the cell surface protein comprises CD80. In someembodiments, the cell surface protein comprises HLADR. In someembodiments, the cell surface protein comprises IgM. In someembodiments, the cell surface protein comprises KTR. In someembodiments, the cell surface protein comprises KLRG1. In someembodiments, the cell surface protein comprises NK1. In someembodiments, the cell surface protein comprises NKg2a. In someembodiments, the cell surface protein comprises TIGIT.

Described herein is a plasmapheresis exchange fluid composition for usein administering plasmapheresis for reducing cellular senescence in anindividual, wherein a total volume of the plasmapheresis exchange fluidcomposition is equal to at least one plasma volume of the individual. Insome embodiments, the composition comprises 5% albumin. In someembodiments, the composition comprises IVIG. In some embodiments, a cellin which the cellular senescence is reduced comprises a lymphocyte. Insome embodiments, the lymphocyte comprises a T-cell. In someembodiments, a cell in which the cellular senescence is reducedcomprises a monocyte. In some embodiments, a cell in which the cellularsenescence is reduced comprises a basophil. In some embodiments, a cellin which the cellular senescence is reduced comprises a neutrophil. Insome embodiments, a cell in which the cellular senescence is reducedcomprises an eosinophil. In some embodiments, a marker associated withthe cellular senescence comprises senescence-associatedbeta-galactosidase (“SA-β-gal”) and wherein the cellular senescence ismeasured by measuring the marker. In some embodiments, theplasmapheresis that is administered is administered over a plurality oftreatment sessions. In some embodiments, two treatment sessions of theplurality of treatment sessions are administered within 72 hours of eachother. In some embodiments, the plasmapheresis that is administered isadministered over a single treatment session. In some embodiments, thecellular senescence is reasured by measuring a marker associated withthe cellular senescence, and wherein expression of the marker ismeasured using flow cytometry. In some embodiments, the expression ofthe marker is measured using a fluorescent conjugated antibody.

BRIEF DESCRIPTION OF THE DRAWINGS

The novel features of the disclosure are set forth with particularity inthe appended claims. A better understanding of the features andadvantages of the present disclosure will be obtained by reference tothe following detailed description that sets forth illustrativeembodiments, in which the principles of the disclosure are utilized, andthe accompanying drawings (also “Figure” and “FIG.” herein) of which:

FIG. 1 is a series of graphs of physical, and mental macro data for afemale collected over the course of six plasmapheresis treatments.

FIG. 2 is a series of graphs of physical, and mental macro data for amale collected over the course of six plasmapheresis treatments.

FIG. 3 is a series of graphs of physical, and mental macro data for afemale collected over the course of six plasmapheresis treatments.

FIG. 4 is a series of graphs of physical, and mental macro data for afemale collected over the course of six plasmapheresis treatments.

FIG. 5 is a series of graphs of physical, and mental macro data for amale collected over the course of six plasmapheresis treatments.

FIG. 6 is a series of graphs of physical, and mental macro data for amale collected over the course of six plasmapheresis treatments.

FIG. 7 is a series of graphs of physical, and mental macro data for amale collected over the course of six plasmapheresis treatments.

FIG. 8 is a series of graphs of physical, and mental macro data for amale collected over the course of six plasmapheresis treatments.

FIG. 9 is a series of graphs of physical, and mental macro data for amale collected over the course of three sham plasmapheresis treatments.

FIG. 10 is a series of graphs of physical, and mental macro data for amale collected over the course of three sham plasmapheresis treatments.

FIG. 11 is a series of graphs of physical, and mental macro data for amale collected over the course of three sham plasmapheresis treatments.

FIG. 12 is a series of graphs of biological aging marker levels measuredin the blood of a female subjected to a regimen of six plasmapheresistreatments.

FIG. 13 is a series of graphs of biological aging marker levels measuredin the blood of a female subjected to a regimen of six plasmapheresistreatments.

FIG. 14 is a series of graphs of biological aging marker levels measuredin the blood of a male subjected to a regimen of six plasmapheresistreatments.

FIG. 15 is a series of graphs of biological aging marker levels measuredin the blood of a male subjected to a regimen of six plasmapheresistreatments.

FIG. 16 is a series of graphs of biological aging marker levels measuredin the blood of a male subjected to a regimen of six plasmapheresistreatments.

FIG. 17 is a series of graphs of biological aging marker levels measuredin the blood of a male subjected to a regimen of six plasmapheresistreatments.

DETAILED DESCRIPTION Aging

Aging coincides with progressions which affect recognizable and oftendeleterious changes in comfort, fitness, appearance and cognition. Whilesome of these progressions manifest as readily identifiable changes inappearance (e.g., in humans, looser skin, increased mouth and nosewidth, and eye droop), the underlying biochemistry—which is believed toinvolve, among other things, complex and multifaceted changes inmolecular and signaling pathways over time—is not yet fully understood.A great deal of research is currently being conducted to betterunderstand the science of aging and changes (e.g. genetic, physiologic)associated with aging at both the micro and macro level of differentorganisms including humans. Generally, changes and conditions that areassociated with aging are considered negative and there is a great dealof benefit in treatments described herein that can improve a healthstatus of an individual by addressing changes and conditions associatedwith aging.

Effects of Aging

Many age-related developments are considered unwelcome and deleteriousto quality of life, such as, for example, hearing and vision loss,arthritis, and loss of cognitive function. This phenomenon is nearlyubiquitous across species, wherein past a certain point, aging coincideswith diminished capabilities and biological function. In addition,certain diseases are highly associated with and possibly interrelatedwith aging, because, for example, aging corresponds with: degenerativeprocesses, diminished recovery or healing capacity, increased propensityfor acute stress and immune response; all of which create an environmentwhere certain disease processes can occur. As an example, rheumatoidarthritis is a disease highly associated with aging, the pathophysiologyof which is associated with tissue degeneration, diminished recovery orhealing and increased propensity for acute stress and immune response.

Physiological effects of aging (i.e. conditions associated with aging)include decreased strength, decreased mobility (and specificallydecreased ability to walk), decreased balance, and subjective changes inmental wellbeing.

Mechanisms of Aging

The mechanisms associated with aging are not at this time fullyunderstood, however, a number of observations provide at least empiricalinsight into factors associated with aging. Observations have, forexample, shown that aging is likely influenced by a number of health andlifestyle factors (i.e. ostensibly non-genetic factors), includingstress, diet, sleep hygiene, and sun exposure. To say that age isinfluenced by these factors is to say that observations seem to indicatethat affecting a change in one or more of these factors can influence arate and/or severity of aging. For example, affecting diet throughmoderate caloric restriction is a well established and reliable meansfor slowing aging which strongly suggests that diet is an importantcomponent of the aging process.

It is also strongly believed, and supported by certain research, thatthere is a genetic component to aging as well, wherein expression ofcertain genes is believed to drive aging related processes and eitherover or under expression of certain genes can lead to slowing oracceleration of the aging process. Gene expression, in the context ofaging, can result in the production of peptides capable of acting on orotherwise affecting targets located relatively remotely, within thebody, from the cell that contains the genes that are expressed. In thisway, genes expressed in one cell type or one tissue type can haveeffects on cells or tissues that are remote from where the gene wasoriginally expressed.

Peptides produced by expression of genes associated with aging may bemodified within the body by other chemical processes which then mayaffect the function of the peptide. Such chemical processes that modifypost-translational peptides include but are not limited to methylation,glycation, and glycosylation. The type of chemical modification as wellas the degree of chemical modification of certain post-translationalpeptides may be both a cause of age-related changes and also a marker ofaging as well. For example, a degree of glycation of a peptide found inblood such as, for example, albumin may directly corelated with aginggenerally or a specific aging process. A degree of chemical modificationof a post-translational peptide can refer to the percent of peptidesfound to have undergone the particular modification and/or a degree ofmodification seen within one or more of the peptides.

Aging is also associated with an increase in the levels ofpro-inflammatory markers in blood and tissues, which is a strong riskfactor for multiple diseases that are highly prevalent and frequentcauses of disability in elderly individuals. This phenomenon is referredto as “inflammaging.” Reducing or even reversing inflammaging in agingpatients is a pathway to treating conditions associated with aging andeven treating or reversing aging itself.

Effect of Aging on Lifespan and Longevity

The term “lifespan” as used herein is the duration of the life of theindividual. Whereas when measured in a population of individualslifespan can be any cumulative measure across the entire population or aportion of the population (i.e. a sub-population), such as, for example,an average lifespan of the population or sub-population, a medianlifespan of the population or sub-population, a variance in lifespan ofthe population or sub-population, and so on.

The term “longevity” as used herein means that an individual orpopulation of individuals has a lifespan or expected lifespan that lastslonger than a reference lifespan. For example, historical data canprovide expected lifespans for a population which can serve as areference lifespan. An “expected lifespan” as used herein may describeany measure of a lifespan of an individual or lifespans within apopulation that can be reasonably used as a predictor or marker for alifespan of another individual or individuals within a population. Forexample, an expected lifespan of an individual having a particularphysiology can be obtained by calculating an average lifespan for apopulation of individuals having the same particular physiology so thatthe average lifespan can serve as a reference lifespan. It should beunderstood that there are a multitude of ways to determine a referencelifespan including using statistical techniques for data of a relevantpopulation such as, for example, mean, median, mode, standard deviation,and variance.

The aging process counters or limits longevity in the sense that aginghas a shortening effect on lifespan. It is well understood that theaging process is not only associated with adverse physiologic change andincreased likelihood of development of life threatening disease, but isalso either a direct or indirect cause of death, which of courseshortens lifespan. It is also well understood that mitigating,preventing, halting, and/or reversing the effects of aging will promoteincreased lifespan and therefore promote longevity. Generally speaking,if you can significantly counter aging, you promote the ability to avoiddeath and therefore live longer, thereby, increasing lifespan andpromoting longevity. This is true for individuals as well as apopulation of individuals. Therefore, aside from aging being a goodtarget for therapy in its own right, therapies that address aging areexpected to promote longevity as well.

Therapies that promote longevity can, therefore, be defined as thosethat promote a relative increase in lifespan in an individual or apopulation and/or therapies that treat, mitigate, and/or prevent theeffects of aging. For example, an expected lifespan for a male humanhaving a particular physiologic feature or features, such as, forexample, dark hair and green eye color, may correspond to 89.4 years,where 89.4 years is the average lifespan of a population of males withdark hair and green eyes. It can then be said that a male human withthese same physiologic features (i.e. dark hair and green eyes)experiences longevity when he outlives the expected lifespan by, forexample, living until the age of 91 years old. Similarly, a populationof males from a particular geographic region, such as, for example,Greece, with these same physiologic features (i.e. a subset of thelarger population of males with dark hair and green eyes but from thespecific geographic region of Greece) can be said to have longevity ifthey all individually have a lifespan that is longer than 89.4 years.Therefore, a therapy associated with or that results in the lifespan ofthe individual or the lifespans of individuals in a population beinglonger than an expected lifespan can be said to be a therapy thatpromotes longevity.

In addition, therapies that promote longevity can also be defined asthose therapies that cause an increase in an expected lifespan of anindividual relative to an existing expected lifespan of a referenceindividual or reference population. For example, a person who is asmoker and has initial expected lifespan, then undergoes a therapy thatcauses him to quit smoking and the quitting of smoking results in alonger expected lifespan.

Biological Age

As used herein, the term “chronological age” refers to the number ofyears that an individual has existed which is a duration of time whichcan be expressed as “age” or “years old”. Chronological age is purely achronological measurement of time having a start point (typically atbirth) and an end point at death and is not determined based on anyproperty of the individual either physical or biological. For example,an individual's chronological age does not change based on how old theirphysical appearance makes them appear nor does it change based on afamily history or genetic feature that would suggest a specific lifespanfor the individual. The term “biological age,” as used herein, on theother hand, is a measure of the aging process and takes into accountphysical, biological, genetic, and biochemical features of anindividual, including but not limited to biological progressions,genetic and epigenetic features, homeostasis measurements, disease-risk,and various molecular changes associated with an individual.

Biological age may be expressed as a duration of years similar to howchronological age is expressed. Biological age may refer to anindividual as a whole or other aspects such as, for example, an organ,an organ system, or other functional system of the individual. Forexample, an individual as a whole may have the biological age of 35 andan immune system age of 27 (i.e. where an immune system age is a subsetor type of biological age).

It is notable that biological age and chronological age may bedecoupled, leading to appearances, energy levels, and/or biologicalprofiles of chronologically older or younger individuals. For example,an individual may be 55 years old (in terms of chronological age) buthave the biological age of 42 years old. Likewise, an individual who is35 years old may have a biological age of 51 years old.

Biological age, at least in some respects, can be thought of as ameasurable performance metric, wherein it is favorable for an individualto have a biological age—as a whole or with respect to a particularfeature of the individual—that is less than the chronological age of theindividual. For example, an individual who has a chronological age of 70years, assuming that they have their own liver and not a transplantedliver, has a liver which has a chronological age of 70 years as well.The same individual of the example may have—based on, for example, oneor more measures discussed above—a biological age of 68 years as anindividual. And, the same individual of the example, may have—based on,for example, one or more measures discussed above—a liver with abiological age of 65 years. That is, in this example an individual mayhave an overall biological age of 68 years whereas an organ of the sameindividual (in this example, their liver) has a biological age of 65years old. In this way biological age can be considered a holisticmeasure or a measure of individual systems and/or a measure of aprocesses within the body of an individual.

In both examples, biological age may be computed using one or moremarkers or factors that correlate with or indicate the biological age ofan individual. Such markers or factors may be measured and/or detectedthrough testing of a biological sample such as, for example, blood,urine, sputum, and sweat.

In addition to population-level variation, biological aging can progressat multiple rates within an individual. Owing to genetics, lifestyle,health, and environmental factors, separate organs, tissue-types, orcells within the individual may exhibit disparate biological ages. Forexample, among a multitude of cumulative and deleterious effects,obesity, diabetes, and renal diseases often accelerate aging in kidneys,such that apparent ages of such a person's kidneys can be much higherthan those of their other organs and omic profiles (e.g., plasmaproteomics). Even in healthy individuals, marked biological agevariation can occur.

While biological aging rates appear to be responsive to ranges ofgenetic and environmental factors, most organisms appear to followinnate and encoded aging timelines. Although biological aging exhibitssome intraspecies variation, upper and lower bounds for aging ratesappear to be primarily determined by species type. For example, whilethere are no known cases of humans living past 125 years of age, bowheadwhales routinely reach 200 years of age, and certain species of clamsconsistently live beyond 500 years. Exemplifying the other extreme,African killifish typically only live for between 4 and 6 months, andexhibit signs of advanced aging as early as 2 months. Supporting agenetic underpinning for aging, a number of human diseases modifybiological aging rates, with Hutchinson-Gilford Syndrome, WernerSyndrome, and Down Syndrome increasing biological aging by about100-800%.

Biological Age Markers

Aging coincides with diverse and complex progressions at molecular,cellular, and tissue levels. As disclosed herein, select aspects ofthese progressions can be monitored to determine biological age in asubject. As many markers for aging can also be responsive to health,lifestyle, and environment, methods for determining biological age canutilize multiple biological markers, and may further use non-ageresponsive biomarkers as calibrants.

Exemplary biomolecules, genetic and epigenetic markers, expressionpatterns, and associated measurement methods which can be useful fordiagnosing chronological and biological age are outlined below. Whilethe biomarkers outlined in this section are of particular utility, theyare intended to serve as examples of age-diagnostic species, and are notintended to be limiting.

Blood-Based Biomarkers

Many of the molecular and biological changes associated with agingmanifest in altered blood composition. At a population level, agingcorrelates with consistent, if nonetheless complex, changes in bloodphenotypes. While some of these changes can be mapped to straightforwardincreases or decreases of single biomarkers, such as progressiveincreases in inflammatory peptide biomarker (e.g., interleukin (IL)-6),C-reactive protein, and tumor necrosis factor-α (TNF-α)) levels withage, aging can also correlate with changes in biomarker processing(e.g., immunoglobulin glycosylation patterns) and ratios among groups ofspecies.

(i) Albumin

For many individuals, albumin, the highest abundance serum protein, canserve as a robust biomarker for aging. Albumin is a family of globulartransport proteins essential for lipid, hormone, and metaboliteclearance and homeostasis. Following typical peak concentrations of 40to 50 mg/mL during late adolescence, serum albumin concentrations oftendecrease by hundreds of μg/mL annually, and exhibiting accelerated ratesof diminution at advanced ages. While a typical serum albumin level isabout 45 and 42 mg/mL for 30-year-old males and females, respectively,by age 60, mean levels decrease to about 42 and 40 mg/mL for males andfemales, respectively.

Furthermore, albumin often exhibits age-dependent structural changeswhich may be useful for aging diagnostics. In most humans, theproportion of glycated albumin increases with age, and typically leadsto diminished function. As albumin activity is essential for multipleforms of homeostasis, the combined impact of diminished albumin levelsand activity can contribute to adverse symptoms of aging (e.g.,diminished energy), and may even augment biological aging rates. Albuminglycation can also evidence other age-related developments, includingdiminished concentrations and functions of regulatory proteins such asinsulin. Accordingly, serum albumin concentration, isoform ratios, andpost-translational modifications (e.g., glycation patterns) can not onlyserve as diagnostic markers for age, but can evidence the severity ofage-related symptoms.

(ii) Ceruloplasmin

For many individuals, changes in ceruloplasmin levels and morphology canbe used to quantitate biological age. Ceruloplasmins are a class ofcopper proteins which participate in iron oxidation and trafficking.Accordingly, ceruloplasmins perform central roles in iron traffickingand reactive oxygen species prevention. Ceruloplasmins exhibitprogressive changes in post-translational modification and isoformpopulations with aging, which can affect activity, localization (e.g.,intravascular versus extravascular distribution), and clearance rate.

While ceruloplasmin consortia typically contain complex arrays ofisoforms and post-translational modification patterns, age-relatedprogressions often manifest as detectable changes in ceruloplasmincopper centers. Such changes can be detected with paramagneticallysensitive spectroscopies such as electron paramagnetic resonance andmagnetic circular dichroism, and can evidence broader changes instructure, isoform ratio, and post-translational modification patterns(e.g., see Musci et al. J Biol Chem, 1993; 268(18):13388-95).Ceruloplasmin also often exhibits age-dependent carbonylation and netcharge, with greater than 3-fold more carbonylation (e.g., as measuredby mass spectrometry) and 0.1 higher isoelectric points (e.g., asmeasured by 2-dimensional gel electrophoresis) in 65-year-old than in15-year-old subjects. Accordingly, ceruloplasmin structure, isoformratios, post translational modification patterns, and combinationsthereof can be used to assess biological age.

(iii) Immunoglobulins

As immunoglobulins are present within blood as complex consortiaspanning varied structural forms, targets, immune activities (e.g.,effector functions and complement binding affinities), and glycosylationpatterns, variations in immunoglobulin populations can serve as strongmarkers for biological aging. Humans express five immunoglobulinisotypes (IgG, IgA, IgM, IgD, and IgE) spanning multiple subclasses(e.g., IgG1, IgG2, IgA1, etc.) and differing in structure,concentration, biodistribution, and immunomodulatory activity. WhileIgG, IgA, and IgM are the second, fifth, and ninth highest abundantproteins in serum, each with mg/mL resting levels, IgD and IgE aretypically present in serum in μg/mL and ng/mL quantities, respectively.

Total immunoglobulin concentrations tend to peak during early adulthood,and then decrease steadily with age. Nonetheless, only someimmunoglobulin isotypes and subclasses exhibit age-dependent changes inserum levels. A recent study (Ritchie et al. J Clin Lab Anal, 1998,12:363-370) identified increases in IgA levels and decreases in IgMlevels with age, as well as age-invariance for total IgG concentrations.However, a follow-on study (Lock and Unsworth. Ann Clin Biochem, 2003;40:143-148) determined that, for certain subjects, only IgG1 and IgG3levels are invariant with age, while IgG2 and potentially IgG4 canexhibit age dependent concentration declines. Contrasting IgA, IgG, andIgM concentration trends, IgD may peak during the first year of life,but remain relatively stable thereafter (Josephs and Buckley, J Pediatr,1980; 96(3): 417-420). For certain subjects, ratios betweenimmunoglobulin isotype and subclass concentrations can provide a strongdiagnostic marker for age. For example, the ratio between IgA and IgM,IgA and IgG2, IgA and IgG4, IgM and IgG2, IgM and IgG4, and/or IgG2 andIgG4 serum levels can evidence age.

Immunoglobulin consortia can also exhibit age-dependent changes inglycosylation. All five human isotypes exhibit diverse glycanmodifications which affect immunomodulatory and biodistributionbehavior. Within each isotype, glycosylation patterns (glycomes) exhibithigh degrees of heterogeneity, as well as health and populationvariance. For example, IgG antibody populations typically exhibitgreater than 30 types of glycans at asparagine 297, in addition tovariable Fab and hinge region glycosylation, some of which vary withdisease status. Nonetheless, age dependent changes in glycosylationpatterns have been observed for all five human isotypes. Within IgGantibody populations, increases in agalactosylation and GlcNAc bisectionand decreases in digalactosylation, sialylation, and afucosylation aretypically observed with aging. Furthermore, there is some evidence thatIgG glycosylation is not only responsive to age, but is a determinantfor the rate of biological aging (Gudelj et al. Cellular Immunology,2018; 333:65-79).

(iv) Glutathione

Glutathione is a versatile biomolecule which participates in oxidativehomeostasis, nitric oxide signaling, aldehyde catabolism, and multipleforms of anabolism. Glutathione is present in micromolar (μM)concentrations in blood as a mixture of reduced monomers and oxidizeddisulfide dimers. The ratio of these two forms is not only responsive toblood conditions, such as reactive oxygen species levels, but shift withage. Augmenting this effect, systemic glutathione levels steadilydiminish with age. As glutathione is critical for mitigating oxidativestress, diminished glutathione levels may be partially responsible forincreased oxidative stress and the progression of stress-relatedconditions (e.g., Parkinson's disease) among elderly individuals.Accordingly, systemic glutathione levels and monomer-dimer ratios canserve as a strong diagnostic marker for biological age. In men andwomen, serum glutathione concentrations steadily diminish from about 1μM at the age of 20 to about 0.5 μM at the age of 60 (Yang et al. JChromatogr B Biomed Appl, 1995; 674(1):23-30).

Physical Function and Appearance

In spite of extensive variation in physical fitness and appearance amongindividuals, diminishing physical abilities and changes in appearanceare universal attributes of aging in humans, including diminishedstrength, diminished ambulation (or walking), and diminished balance.

Methods for Measuring Biological Age

The present disclosure provides a range of methods for ascertainingbiological age and biological aging rates with biomarker analysis.Underlying the molecular complexity of aging, a number of species changein predictable manners during aging, and therefore provide metrics forbiological and chronological age. As an individual ages, certainbiomarkers can include an increase or decrease in concentration, analteration in state (e.g., glycation of albumin andmethylation/demethylation of genomic DNA), a change in form (e.g.,isoform ratios of a particular protein), a change in activity, or acombination thereof. Accordingly, as detailed further herein, assessmentof one or more of these biomarkers (e.g., identifying concentration,state, form, and/or activity) can be used to determine biological age.

While some biological age measurements can identify a biological age ofan individual, others identify biological ages of individual cells,tissues, organs, or systems (e.g., immune or endocrine systems). In manyindividuals, biological aging progresses in cell-, tissue-, organ-,and/or system-specific manners, reflecting distinct environments,stresses, and genetic and regulatory architectures. In the absence oftrauma or aberrant health, the range of biological ages of anindividual's tissues and cells is often small, for example less than agemeasurement experimental error. However, many individuals exhibitmultiple, disparate ages. For example, relative to their chronologicalage, an individual may have a young brain age and advanced immune age.For such an individual, biological age may reflect a sort of median ofcell, tissue, organ, and/or system-specific biological ages.Alternatively, the biological age of the individual may be expressed asa set of distinct, organ or system specific biological ages.

A biological age measurement can utilize one biological age marker or aplurality of biological age markers. In many cases, the accuracy of abiological age determination method increases as more age markers areutilized for analysis. However, for many individuals, use of a singleage marker or a small set of age markers are sufficient for accuratelydetermining biological age and/or biological aging rate, for examplewith a standard error of less than 7 years, less than 5 years, or lessthan 3 years. Examples of methods for assessing biological age areprovided in TABLE I.

A method for biological age determination can utilize a single assay ora plurality of assays from TABLE I, a cytokine inflammatory markerpanel, a metabolomic assay, peripheral blood mononuclear cell (PBMC)analysis, genomic methylation analysis, inflammatory marker analysis, ora combination thereof. The assay or plurality of assays can assessholistic biological age, organ and/or system-specific biological age, ora combination thereof. The biological age(s) determined for a subjectcan be used to calibrate a treatment, such as a treatment for agingdisclosed herein.

The assay or plurality of assays can be performed at regular intervals,for example once per month, once every three months, once every sixmonths, or once per year. In this way (as well as with agingrate-diagnostic methods), biological age(s) determined from the assay orplurality of assays can also be used to determine biological aging ratein the subject, for example to determine whether an aging treatment isslowing a rate of aging in a subject, to calibrate a treatment method toachieve a target age or aging rate in a subject, to decouple diseasemarkers from aging-related symptoms (e.g., to determine whether raisedHbA1c levels stem from disease or aging), or a combination thereof.

TABLE I Exemplary Age-Measurement Assays Antibody Assays AntinuclearAntibody (ANA) Screen (IFA) with Reflex to Titer Assay Rheumatoid FactorAssay Thyroid Peroxidase Antibody (Anti-TPO) Assay QuantitativeImmunoglobulin Assay Proteomic Assays Fibrinogen Assay Creatinine KinaseAssay Hemoglobin A1C Assay Metabolomic Assays Cholesterol, Direct LDLAssay HDL Cholesterol Assay Total Cholesterol Assay Blood Glucose AssayOther Urine Analysis

Antibody Assays

A method for determining biological age can include an assessment ofantibody concentration, type, and structure. While antibodies arepresent as complex consortia spanning multiple isotypes (in humans, IgA,IgD, IgE, IgG, and IgM), paratope structures, and processing (e.g.,glycanation), changes among these consortia can be diagnostic ofbiological aging. For example, as further detailed herein, ratios ofantibody isotypes typically shift with aging. Furthermore, individualantibody types, such as anti-nuclear and anti-thyroid peroxidaseantibodies, change in concentration with age, and can thereby serve asmarkers for aging. A number of illustrative antibody assays are outlinedbelow. It is contemplated herein that additional antibody assays may beused with methods of the present disclosure.

(i) Anti-Nuclear Antibody Screen

An anti-nuclear antibody (ANA) screen measures cell nucleus-bindingantibody concentrations in blood, plasma, or serum. While a range of ANAsubtypes are present in humans, most ANA screens measure total ANAantibody concentration, most commonly with indirect immunofluorescenceand enzyme-linked immunosorbent (ELISA) detection following cell binding(e.g., to HEp-2 cells). ANAs are associated with a range of disorders,many of which are associated with aging. However, even among healthyindividuals, ANA blood concentrations tend to increase with age, withelderly individuals often exhibiting 3- or greater-fold ANA levelsrelative to younger individuals (Xavier et al. Mech Aging Dev, 1995;78(2):145-54). Accordingly, a method consistent with the presentdisclosure can utilize a blood ANA concentration measurement todetermine biological age or biological aging rate.

(ii) Rheumatoid Factor Assay

An age-diagnostic method can assess blood (e.g., whole blood, serum,plasma) levels of rheumatoid factor (RF) factor, an autoantibody againstthe Fc portion of IgG and implicated in a number of age-relatedconditions, including rheumatoid arthritis and diminished bone density.While rheumatoid factor can present as a combination of immunoglobulinisotypes (e.g., IgA, IgD, IgE, IgG, and IgM) with ranges of Fc epitopesand binding affinities, total rheumatoid factor levels can be identifiedwith a number of binding assays, including indirect immunofluorescenceand enzyme-linked immunosorbent (ELISA). In most people, Rheumatoidfactor typically appears between the age of 30 and 70 and progressivelyincreases in concentration with age. Accordingly, a method for measuringbiological age consistent with the present disclosure can includemeasuring blood RF concentration.

(iii) Thyroid Peroxidase Antibody Assay

A thyroid peroxidase antibody assay measures concentration ofautoantibodies which target thyroid peroxidase (TPO), an enzymeessential for thyroid hormone production. As thyroid peroxidase is thetypically the most prevalent thyroid autoantigen, thyroid peroxidaseantibody levels can be reflective of total anti-thyroid antibody levels.While thyroid antibodies (including thyroid peroxidase antibodies) areimplicated in a number of diseases, thyroid antibody levels also tend toincrease spontaneously with age (Chen et al. Endocrinology, 2010;151(9):4583-4593), thereby allowing them to serve as markers forbiological aging. Accordingly, a method of the present disclosure canutilize blood thyroid antibody and/or blood thyroid peroxidase antibodylevels to determine biological age. Furthermore, in some subjects, thetargets of thyroid peroxidase antibodies shift with age, withanti-domain A antibodies typically increasing in prevalence (Czarnockaet al. Clin Endocrinol (Oxf), 1998; 48(6):803-8).

(iv) Quantitative Immunoglobulin Assay

A quantitative immunoglobulin assay can assess total antibody levels ina subject sample. Typically, quantitative immunoglobulin assays measuretotal antibody levels in blood. However, some assays assessconcentration by antibody isotype or subtype. For example, aquantitative immunoglobulin assay may measure total IgG and IgAconcentrations, or may separately determine concentrations of individualantibody subtypes (e.g., IgG1, IgG2, IgA1, IgA2, etc.). As in manysubjects, total antibody, antibody isotype, and antibody subtypeconcentrations change with age (Crisp and Quinn. Allergy Asthma Proc,2009; 30(6):649-54), quantitative immunoglobulin assays can be used todetermine biological age.

(v) Glycanation Assays

Biological aging can be evidenced by immunoglobulin glycosylationprofiles. All five human antibody isotypes exhibit glycosylationpatterns. While the positions of glycosylation are partially isotypedependent, the types of oligosaccharides, or glycans, which couple atthese positions can affect antibody localization, subcellularpartitioning, aggregation, and Fc and complement receptor affinities.Furthermore, these glycation patterns can be reflective of age,environment, and health status. For example, IgG galactosylation oftendecreases with age, while fucosylation, sialylation, and bisection canrespond to age in a gender-specific manner (Gudelj et al. CellularImmunology, 2018; 333:65-79). Accordingly, a method can utilize anantibody glycanation profile to assess biological age.

In addition, a glycanation assay for use in assessing biological age iscurrently marketed as the Glycanage test.

Proteomic Assays

Humans are estimated to have between 20 thousand and 5 million proteins,depending in part on structural variant classifications (e.g., whethersplicing variants constitute distinct proteins), such that proteomicshifts with aging and changes in health status are often complex.Nonetheless, as proteins participate in and regulate the majority ofbiological processes, changes in physiology, including those associatedwith aging, are often reflected in protein expression and activity. Tothis point, changes in the human proteome with age are thought toinclude both drivers of (e.g., diminished superoxide dismutase andcatalase activity) and responses to (e.g., increased fibrinogenconcentration) biological aging. Changes in the human proteome with ageinclude up- and downregulation of individual proteins, as well asproteome-wide changes in abundances, ratios, and activity levels.Despite the complexity of the human proteome, a number of proteins areknown to change with aging in detectable manners. Accordingly, a methodfor determining biological age can include measuring the abundance,state, distribution, modification, or activity of an individual proteinor collection of proteins. In many such cases, the method includesmeasuring the concentrations of a small number of blood proteins.

However, a method for determining biological age can also take a broad,proteomic view to assess biological age. As human blood contains over5000 types of proteins, aggregate analysis of tens, hundreds, orthousands of proteins can often correlate small proteomic shifts tobiological age, even in the absence of statistically significantindividual protein biomarkers. Advances in high-throughput proteomicanalysis, such as in liquid chromatography-mass spectrometry, proteinsequencing, and multiplexed immunoassays, can enable rapidquantification of hundreds or thousands of proteins from individualsamples.

(i) Fibrinogen Assays

Fibrinogen activity and blood level often changes with age. Fibrinogenis blood-based glycoprotein complex which polymerizes to fibrin tofacilitate blood clotting. While fibrinogen levels can be responsive tohealth and inflammation, baseline fibrinogen levels typically increasewith age, rising by about 250 μg/mL per decade, or from about 2.2 mg/mLto about 3.2 mg/mL from the age of 25 to the age of 65 in manyindividuals (Hager et al. Aging (Milano), 1994; 6(2):133-8). As elevatedfibrinogen levels are associated with a number of age-relatedconditions, including increased cardiovascular disease susceptibilitiesand diminished kidney and liver function, fibrinogen levels oftencorrelate with many recognizable, qualitative aspects of aging.

Fibrinogen assays typically assess at least one of fibrinogen activityand concentration. Fibrinogen activity is typically measured indirectlywith blood clotting tests, such as thrombin and prothrombin time tests,thromboelastometry, and qualitative clotting assays. However, theseassays can have limited ability to distinguish between low fibrinogenlevels and diminished fibrinogen activity. Alternatively or in additionto activity analyses, some assays directly measure blood fibrinogenconcentration, with a wide range of immunoassays commercially availablefor such measurements.

(ii) Creatinine Kinase Assays

Changes in the creatine system, which furbishes muscle cells withphosphocreatine for rapid energy production, can reflect aging, withcreatine and associated metabolite levels typically decreasing withaging. Nonetheless, creatinine, the primary breakdown product ofphosphocreatine catabolism, typically increases in blood concentrationwith age. This discrepancy is likely due to diminished capacity forcreatinine recycling and clearance. Creatinine kinase, an intramuscularenzyme which converts creatinine back into phosphocreatine for furtheruse, diminishes in concentration with age, leading to higher levels increatinine in blood, muscles, and some extravascular spaces.Accordingly, blood and muscle creatinine kinase levels can be effectivefor determining biological age.

(iii) Hemoglobin A1c Assays

During circulation, hemoglobin can promiscuously couple to blood glucosein a process referred to as glycation. While the baseline rate forglycation is typically low, hyperglycemia, hormonal insensitivities, andaging can enhance glycation rate, leading to higher concentrations ofglycated hemoglobin. One of the most prevalent forms of the resultantglycated hemoglobin is HbA1c (also referred to as hemoglobin A1c), whichcontains glucose attached at one or both (3-peptide N-terminal valines.While high levels of HbA1c are most commonly ascribed to hyperglycemiain diabetes, changes in glucose tolerance and glycemic regulation withaging tend to increase HbA1c baselines irrespective of health status,leading to age-related shifts in HbA1c populations which can be small(for example 5.4% to 5.6% from the age of 25 to greater than 65), butnonetheless significant. When corrected for health and environmentalfactors, HbA1c levels can be diagnostic for biological age (Masuch etal. BMC Endocrine Disorders, 2019; 19, 20). HbA1c can be measured with avariety of techniques, including quantitative chromatography (e.g., byhigh-performance liquid chromatography), immunoassays, and capillaryelectrophoresis.

(iv) Cytokine and Inflammatory Marker Panels

A method for assessing biological age can include analysis of one ormore cytokines. In many cases, the biological age assessment includesdetermination of one or more blood cytokine concentrations. Oftenreferred to as inflammaging, aging often coincides with increased bloodcytokine levels through increased cytokine production and diminishedanti-inflammatory responses. Central to this process, a number ofpro-inflammatory markers, such as C-reactive protein and serum amyloidA, increase by multiple-fold levels through middle and advanced age,often leading to an imbalance between pro- and anti-inflammatorycytokines. As these changes in cytokine levels often foster chronicinflammation, weakened immune states, and diminished energy metabolism(Salvioli et al. Current Pharmaceutical Design, 2006; 12:3161-71),cytokine imbalance is likely both a cause and effect of aging.Accordingly, blood cytokine concentrations can be correlated withprogressive age-related changes in cytokine levels to assess biologicalage.

A method for determining biological age can include measuring bloodconcentrations of one or more cytokines. While a blood concentration ofa single cytokine can be sufficient for determining biological age, inmany cases the method includes measuring blood concentrations ofmultiple cytokines. As non-limiting examples, a method for determiningbiological age can assess levels of one or more of C-reactive protein,soluble tumor necrosis factor receptor 1 (sTNF1), soluble tumor necrosisfactor receptor 2 (sTNF2), tumor necrosis factor α (TNF-α),interleukin-1 α (IL-1α), interleukin-1 β (IL-1β), interleukin-6 (IL-6),and interleukin-10 (IL-10), all of which typically increase inconcentration with age (Salvioli et al.; Glossop et al. ArthritisResearch & Therapy, 2005; 7:R1227). Among these cytokines, C-reativeprotein, TNF-α, IL-1α, IL-10, IL-6, and IL-10 are implicated inincreased proinflammatory responses with aging, and may play roles inatherosclerosis, and insulin insensitivity, among other conditions(Salvioli et al.), while sTNF1 and sTNF2 likely aggravate arthritis(Glossop et al.). Accordingly, blood concentrations of one or morecytokines can evidence aging and age-related symptoms.

In addition, a test for inflammatory changes in the context ofbiological age (i.e. an inflammatory age test) is currently marketed asthe iAge test.

Expression of certain cell surface proteins may be associated withinflammatory processes as well including CD16, CD25, CD27, CD38, CD57,CD80, HLADR, IgM, KIR, KLRG1, NK1, NKg2a, and TIGIT. These cell surfaceproteins may be measured and quantified using flow cytometry.

Metabolomic Assays

As metabolism encompasses the biochemistry of growth, energy productionand consumption, and certain forms of signaling, age-related changes areoften reflected by metabolomic shifts. As with proteomic analysis,metabolomic profiling can query individual biomolecules which areresponsive to aging, can broadly profile portions of a subject'smetabolome (e.g., by profiling tens, hundreds, or thousands ofmetabolomic biomarkers), or can include a combination thereof. Owing toa higher number of available metabolites and lower variance than celland tissue profiling, metabolomic analysis often focuses on bloodmetabolites. As over 18000 metabolites have been identified in humanblood (Adav and Wang. Aging Dis, 2021; 12(2):646-661), a biological agemeasurement could, in theory, accurately determine biological age with abroad, non-targeted profiling approach, even in the absence of a strongdiagnostic aging marker. However, a number of age diagnostic studieshave identified robust metabolite biomarkers, which, alone or incombination with other measurements, can accurately measure biologicalage.

(i) Total Cholesterol Assays

Cholesterol is complexed in a variety of lipid-protein macromolecularstructures, commonly referred to as lipoproteins, for transport throughthe blood. Although lipoproteins differ in terms of a number ofproperties, including size, composition, and receptor affinities, inhumans, lipoproteins are divided into five major classes based primarilyby on densities. High-density lipoproteins (HDL) tend to have the lowestvolumes and highest protein and phospholipid contents of the fiveclasses of lipoproteins, with diameters typically ranging from 5-15 nm,densities of greater than 1.063 g/mL, and protein accounting forapproximately ⅓ of their mass. Low-density lipoproteins (LDL) tend tohave slightly lower masses of between about 1.019 and 1.063 g/mL,slightly larger diameters of about 18-28 nm, and higher cholesterolcontent approaching nearly 50% (by mass). Intermediate-densitylipoproteins (IDL), tend to have densities of between 1.006 and 1.019g/mL, similar cholesterol content as high-density lipoproteins, anddiameters ranging from 25 to 50 nm. Very low-density lipoproteins (VLDL)are usually characterized as having densities of between 0.95 and 1.006g/mL, relatively low protein content (typically about 10% by mass), anddiameters of between about 30 and 80 nm. Finally, chylomicrons, thelargest lipoproteins, typically have densities below 0.95 g/mL, proteincontent below 2% (by mass), and diameters ranging from 75 to 1200 nm.

Cholesterol analysis often distinguishes between at least somelipoprotein types when assessing cholesterol levels. For example, lipidpanels typically distinguish between HDL and LDL-bound cholesterol, andsometimes further distinguish VLDL, IDL, and chylomicron cholesterolcontent. However, for many conditions, total blood cholesterol contentis sufficient for accurate diagnosis.

In particular, for some individuals, total cholesterol content can beindicative of biological age. In men, blood cholesterol levels tend tosteadily increase from about 1.6 mg/mL at the age of 18 to about 2 mg/mLat the age of 50, after which time blood cholesterol levels tend to dropby about 0.05 mg/mL every decade. Slightly more punctuated trends tendto be observed in women, with blood cholesterol levels typicallyincreasing from about 1.7 to 2.1 mg/mL between the ages of 18 and 60,and then plateauing or declining at a slower rate than in men throughadvanced aging (Yi et al. Scientific Reports, 2019; 9:1596).Accordingly, a method disclosed herein can use blood cholesterol levelto determine biological age.

A number of enzymatic, chemical, electrochemical, and spectroscopicmethods can be used to determine cholesterol levels. Commonly,cholesterol, either pooled or collected from a lipoprotein fraction, iscoupled to a chromophore or fluorophore through its C3-hydroxyl, andquantitated spectrophotometrically (Li et al. Journal of Food and DrugAnalysis, 2019; 27(2):375-386).

(ii) Cholesterol, Direct LDL Assays

In many individuals, low-density lipoprotein (LDL), one of the primarycarriers for cholesterol in blood, increases in concentration throughmiddle age, and then remains stable or decrease with advanced aging(McAuley and Mooney. Med Hypotheses, 2017; 104:15-19). In men, LDL tendsto plateau between 50 and 60 years of age, whereas in women, this trendis prolonged, with maximum concentrations typically occurring between 60and 70 years of age (Kreisberg and Kasim, 1987; 82(1): 54-60). Althoughoften requiring corrections for certain factors, such as estrogen levelsin women, LDL levels can be a powerful metric for biological age.Accordingly, a method for determining biological age can includedetermining an LDL level of a subject.

Often, LDL is simultaneously measured along with other lipids andlipoproteins as part of lipid profiles or panels. In some panels, LDL isindirectly identified through total cholesterol measurements. An LDLmeasurement can identify LDL, LDL-cholesterol (LDL-C, the amount ofcholesterol contained within low-density lipoproteins), or both. Ascholesterol is primarily carried within LDL, high-density lipoproteins(HDL), and very low-density lipoproteins (VLDL), LDL levels are oftendetermined without measuring LDL, but rather by subtracting non-LDLabundances from a measured level of cholesterol. For example, lipidpanels in the United States commonly estimate LDL levels by subtractingmeasured HDL and triglyceride levels from measured total bloodcholesterol.

LDL can also be measured directly. In many cases, these methods involveseparation of LDL from other lipoproteins and lipids, including HDL,VLDL, and lipoprotein a (Lp(a)). Common methods for achieving theseseparations include centrifugation (e.g., ultracentrifugation) which canseparate lipoproteins and lipids by density; electrophoresis, which canseparate lipoproteins and lipids based on size and charge;precipitation, in which lipoproteins and/or lipids are selectively drawnfrom solution; by homogeneous methods, which utilize combinations ofconditions, binding molecules, and polymers to separate lipoproteinfractions; and by combinations thereof. The separated lipoproteins andlipids can then be quantified with a range of enzymatic,electrochemical, chemical, and spectroscopic methods (Nauck et al.Clinical Chemistry, 2002; 48(2):236-254).

(iii) Cholesterol, Direct HDL Assays

Similar to LDL, HDL levels tend to decrease in concentration withadvanced age. Complicating the relationship between HDL and aging, lowHDL may correlate with mortality, potentially obfuscating otherwisereliable trends in lowered HDL abundance with age (Walter,Arteriosclerosis, Thrombosis, and Vascular Biology, 2009; 29:1244-1250).Nonetheless, a number of studies have defined clear decreases in HDL asa function of age, especially among males (Ferrara et al. Circulation,1997; 96:37-43). Accordingly, HDL levels, alone or in combination withother cholesterol data (e.g., LDL, total cholesterol, etc.), can be usedto assess biological age.

As with LDL-cholesterol, HDL-cholesterol is typically measured throughseparation followed by cholesterol quantification. However, a number ofmethods (in particular some homogeneous methods) allow HDL-cholesterolto be measured simultaneously with other forms of cholesterol.

(iv) Blood Glucose Assays

In many individuals, aging coincides with increased resting andpost-meal glucose levels. As blood glucose levels are responsive tomultiple age-sensitive regulatory mechanisms, including insulin andincretin sensitivities, and can contribute to age-related developments,such as increase HbA1c and albumin glycation, blood glucose levels cancapture a broad spectrum of aging-related progressions, and can serve asreliable diagnostic markers for aging. In men and women, resting andpost-meal glucose levels tend to increase at rates of about 7 to 11μg/mL per decade, while 2-hour-post-meal levels tend to increase by amore pronounced rate of 56-66 μg/mL per decade (Chia et al. CirculationResearch, 2018; 123(7):886-904). Following from these trends, bloodglucose levels (either resting, post-meal, or a combination thereof) canbe used to determine biological age.

A number of cheap methods are available for blood glucose measurement.Most commonly, blood glucose is detected through enzymatic (e.g.,through hydrogen peroxide generation by glucose oxidase) or chemicaloxidation.

Urine Analysis

While water, urea, and sodium chloride account for greater than 96% ofits mass, urine contains a complex mixture of chemicals which canacutely reflect age, health and environment. Over 450 species ofmicrobiota and 3000 molecules have been identified in urine, of which480 have not been detected in blood (Bouatra et al. PLoS One, 2013;8(9):e73076). Many of the molecules in urine are waste materials,including biproducts of metabolism and damaged and solubilizedbiomolecules. Accordingly, age-related changes within a subject, inparticular their kidney, liver, and bladder ages, are often reflected inurine composition (Harpole et al. Expert Rev Proteomics, 2016;13(6):609-626).

Peripheral Blood Mononuclear Cell Analysis

A method for determining biological age can include peripheral bloodmononuclear cell (PBMC) analysis. PBMCs include blood cells with roundnuclei, such as T lymphocytes, B lymphocytes, natural killer cells, andmonocytes (and contrasting non-nucleated cells, such as erythrocytes,and cells with monolobed nuclei such as granulocytes). A number of PBMCphenotypic distributions have been shown to shift with age. In certainindividuals, T cell populations shift from CD28+ towards CD95+ with age,suggesting reduced proliferation and increase apoptosis (Li et al. J IntMed Res, 2020; 48(7)). Furthermore, aging often coincides with increasedmonocyte and regulatory T cell and decreased naïve T cell populations(Huang et al. PNAS, 2021; 118(33):e2023216118). Accordingly, PBMCpopulations and subpopulation polarization can be used for biologicalage analysis. Such analysis may include PBMC immunophenotyping, forexample to identify T lymphocytes, B lymphocytes, and natural killercells, as well as CD4, CD8, CD28, naïve, effector, and central memorycell subpopulations. As an illustrative example, PBMCs can be isolatedfrom whole blood, and characterized with flow cytometry, for example asoutlined in Li et al.

Flow cytometry may also be combined with fluorescent conjugatedantibodies so that in addition to identifying certain cell types, cellsurface proteins may be identified and/or quantified as well. Types ofcell surface markers (i.e. proteins located on a cell surface) found oncells in blood that may be identified and quantified using flowcytometry include CD16, CD25, CD27, CD38, CD57, CD80, HLADR, IgM, KIR,KLRG1, NK1, NKg2a, and TIGIT. Types of cells that may have these markersinclude white cells and specifically lymphocytes, neutrophils,monocytes, basophils, and eosinophils. For example, T lymphocytes mayinclude one or more of cell surface markers that may be identifiedand/or quantified using flow cytometry.

Cellular Senescence Assay

A biological age measurement can include senescent cell populationanalysis. While cellular senescence is characterized by arrested growthand termination of cell division, senescence coincides with changes incellular phenotypes. Senescent phenotypes are not only adverse forcellular function, generally inducing lower levels of fitness (e.g.,diminished energy metabolism and elevated oxidative stress), but alsotend to coincide with deleterious secretory behavior, suchproinflammatory exosome production, and senescence-associated secretoryphenotype (SASP) secretion behavior. Exemplifying this point, a recentmouse model experiment demonstrated that as few as 1/10,000 senescentcells in healthy adult mice are sufficient for causing systemicdisfunction, metabolic stress, and accelerated aging (Xu et al. Nat Med,2018; 24(8):1246-1256). Accordingly, cellular senescence is not onlyharmful to senescent cells, but can accelerate aging and aggravate agingsymptoms in disparate cells within an organism.

Senescence phenotypes typically affect detectable changes in expressionprofile. At a cellular level, non-terminally differentiated senescentcells can often by identified by their inability to replicate or undergoDNA synthesis. On a genetic level, senescent cells can also typically beidentified based on their altered regulation of proliferation-associatedand growth inhibitory genes (Itahana et al. Methods to Detect Biomarkersof Cellular Senescence. In Methods in Molecular Biology: BiologicalAging: Methods and Protocols. Jumana Press Inc.). However, senescentcells are most commonly identified by senescence-associated markers,which can include upregulation of the lysosome-associated proteinβ-galactosidase (Dimri et al. Proc. Natl. Acad. Sci, 1995; 92:9363-9367); the DNA damage response marker H2AX (Campisi, J. Annu. Rev.Physiol, 2013 75:685-705); tumor suppressors p16ink4a, p21, and p53(Rufini et al. Oncogene, 2013; 32: 5129-5143); and altered secretoryprofiles (for example increased proinflammatory cytokine production,Itahana et al.).

A marker associated with the cellular senescence comprisessenescence-associated beta-galactosidase (“SA-β-gal”) which can bemeasured in blood and used as a measure of cellular senescence.

Genomic Methylation Assays

DNA methylation is a prevalent epigenetic modification which canstrongly affect expression, and correspondingly phenotype, in a subject.Recent studies have demonstrated that aging coincides with genome-wideDNA methylation and demethylation, with a greater prevalence ofdemethylation than methylation during most human lifespans. While totalgenomic methylation can correlate with age, a number of recent studieshave identified specific sites which can serve as aging markers based onmethylation or hydroxymethylation status (Salameh et al. Front. Genet.,2020; 10). Accordingly, a method of the present disclosure may determinemethylation status at a site or plurality of sites in genomic DNA toascertain biological age.

Inflammatory Marker Analysis

In some cases, inflammatory markers can be used to assess biological agein a subject. As used herein, an inflammatory marker can be a specieswhich causes or increases in concentration in response to inflammation.Inflammation tends to increase with age, often diminishing immunefunction, contributing to frailty, and augmenting conditions such asarthritis, asthma, atherosclerosis, and certain forms of dementia.Nonetheless, only some inflammation-related markers increase inconcentration with age. For example, a recent study identified Chemokine(C-X-C motif) ligand 9 (CXCL9), EOTAXIN, macrophage inflammatory protein(Mip-1α), LEPTIN, IL-1β, interleukin-5 (TL-5), interferon-α (IFN-α),interleukin-4 (IL-4), TNF-related apoptosis-inducing ligand (TRAIL),interferon-7 (IFN-7), Chemokine (C-X-C motif) ligand 1 (CXCL1),interleukin-2 (TL-2), transforming growth factor-α (TGF-α), plasminogenactivator inhibitor (PAI)-1, and leukemia inhibitory factor (LIF) aspotential biomarkers for inflammation, while other inflammatory responsemarkers, such as TL-6 and TNF-α, have minimal association with aging(Sayed et al. Nature Aging, 2021; 1: 598-615). Stemming from this typeof observation, a method of the present disclosure can utilize one ormore inflammatory markers to determine biological age.

Safety Analysis

An aging treatment disclosed herein can be combined with a safety orhealth assessment. In addition or alternatively to monitoring biologicalage in a subject receiving treatment for aging, one or more markers forhealth can be monitored to ensure treatment efficacy, to provide acalibrant for age measurement, or a combination thereof. Some agingtreatments can cause adverse effects, such as diminished red blood cell(RBC) count or diminished liver function in certain subjects. Monitoringhealth prior to or concurrently with an aging treatment can allow thetreatment to be tailored to the subject, can determine whether a subjectis suitable for an aging treatment, and can ensure that the treatment ismodified or ceased following an adverse response.

Furthermore, in some cases, markers for overall health can be importantfor determining biological age. As many age markers fluctuate inresponse to health status, calibration to a subject's health can berequisite for accurate biological age assessments. For example, certainconditions can increase autoantibody concentrations by greater degreesthan aging, rendering such autoantibody analyses effective for onlycertain subjects. Nonlimiting examples of health assessments areoutlined in TABLE II. A subject of an aging treatment can be assessedwith one or with a plurality of health assessments outlined in TABLE IIor otherwise consistent with methods disclosed herein.

TABLE II Complete Blood Count (CBC) Assay Total Protein Assay LiverFunction Assay Blood Urea Nitrogen (BUN) Assay Creatinine AssayC-Reactive Protein (CRP) Assay

(i) Blood-Based Assays

A subject of an aging treatment method, before, during, and/or followingthe treatment, can be assessed with a complete blood count (CBC) assay.Complete blood count assays typically quantitatively measure multiplecomponents of blood, such as red blood cells, white blood cells,hemoglobin and platelets, as well as health and aging factors, such asred blood cell to plasma ratios. In addition to screening for overallhealth, complete blood count assays can identify adverse responses tosome aging treatments, such as blood diffusion-based anemia.

Analogously, a subject of an aging treatment can be assessed with atotal protein test to determine protein levels in a biofluid (e.g.,blood). While some total protein tests quantify specific proteins, suchas albumin and globulin concentrations and/or ratios (e.g., withimmunoassays), others determine total protein concentration in thebiofluid (e.g., based on the 280 nm protein band in a spectrophotometricabsorbance assay). Total protein tests can identify some potential sideeffects of aging treatments, including fatigue, edema, and nutritionaldeficits.

Liver function assays can test for a number of substances indicative ofproper liver function and health. Many liver function assays determineblood levels for liver-based or secreted enzymes, including alaninetransaminase, aspartate transaminase, alkaline phosphatase, albumin, andgamma-glutamyltranserase. A liver function assay can also assess levelsof metabolites regulated (e.g., cleared) by the liver, such asbilirubin, a major heme degradation product. A liver function assay canalso determine a quality of blood, such as pH or prothrombin time (howquickly blood clots). A specific set of assessments included in a liverfunction assay can depend on the health of the subject, as well as thetype of aging treatments and diagnostics to which they are subjected.

A blood urea nitrogen (BUN) assay can assess kidney and metabolicfunction in a subject receiving an aging treatment. Blood urea nitrogenassays measure urea levels in blood. Blood-based ureas, which primarilyderive from protein degradation, are maintained at low levels by kidneyfiltration. In addition to improper kidney function, high blood urealevels can indicate dehydration (a risk associated with some blooddilution methods), internal bleeding, and shock.

Creatinine assays provide an additional form of assessment for kidneyfunction. Creatinine assays measure concentrations of creatinine, acatabolic waste product, in blood. While kidneys typically filter andthus maintain low levels of creatinine in blood, this function ishindered by a number of conditions which are identifiable withcreatinine assays. In certain subjects, use of a creatinine assay can beimportant for monitoring kidney health prior to, during, or following anaging treatment.

C-reactive protein (CRP) assays can serve as a measure for inflammationand infections in a subject of an aging treatment. As some agingtreatments increase susceptibility to infection, C-reactive proteinassays can be important measures for aging treatment efficacy andsubject health.

Methods for Performing Plasmapheresis Treating and Preventing theEffects of Aging Using Plasmapheresis

The term “plasmapheresis” as used herein is interchangeable with theterm “therapeutic plasma exchange,” and plasmapheresis is a form ofapheresis wherein some amount of a plasma of an individual is withdrawnfrom the body of the individual and removed. Plasmapheresis methods aredescribed herein for treating and/or preventing a symptom or a conditionassociated with aging. In treating and/or preventing a symptom or acondition associated with aging, the plasmapheresis methods describedherein may also increase lifespan and promote longevity.

A plasmapheresis treatment typically comprises—and begins with—thewithdrawing of whole blood from an individual receiving theplasmapheresis treatment. Whole blood that is withdrawn is separatedinto a cellular fraction and a plasma fraction. The term “cellularfraction” as used herein can refer to or comprise red blood cells, whiteblood cells, and platelets. The terms “plasma fraction” or “plasma” asused herein can refer to or comprise a liquid portion of whole bloodwhich contains, among other things, proteins, electrolytes, vitamins,and hormones. Typically, in a plasmapheresis treatment, the plasmafraction that is separated is removed while the cellular fraction isreturned to the individual receiving the plasmapheresis treatment. Asused herein in the context of plasmapheresis administration (or anyother type of apheresis procedure), the terms “withdraw,” “withdrawal,”“withdrawn,” and “withdrawing” (or any other conjugation of “withdraw”)means to draw blood out (actively or passively) from the vascular systemof an individual receiving plasmapheresis (or other type of apheresisprocedure) which may be achieved using any suitable vascular access,which includes but is not limited to peripheral intravenous lines andcentral lines. As used herein in the context of plasmapheresisadministration (or any other type of apheresis procedure), the terms“return” or “returning” (or any other conjugation of “return”) or“infuse,” or “infusing” (or any other conjugation of “infuse”) means toreturn blood back (actively or passively) to the vascular system of anindividual receiving plasmapheresis (or other type of apheresisprocedure) which may be achieved using any suitable vascular access. Asused herein in the context of plasmapheresis administration (or anyother type of apheresis procedure), the terms “remove,” “removal,”“removed,” and “removing” (or any other conjugations of “remove”) meansto remove at least a portion of whole blood withdrawn from an individualreceiving plasmapheresis (or any other apheresis procedure) and notreturning the at least portion of the whole blood to the individualreceiving plasmapheresis so that it is removed from their body. As usedherein, in the context of plasmapheresis (or any other apheresisprocedure), the terms “separate,” “separated,” or “separating” (or anyother conjugation of “separate”) means to separate apart components ofblood from one another. For example, in plasmapheresis, whole blood iswithdrawn and plasma is separated from the cellular fraction of thewithdrawn whole blood. As used herein, the term “plasmapheresis” may becombined with other terms such as “therapy” (i.e. plasmapheresistherapy) or “treatment” (i.e. plasmapheresis treatment) or “procedure”(i.e. plasmapheresis procedure) and, unless otherwise indicated, nospecific meaning should be attributed to the use of one of these termsor the other in the context in which they appear. Of note, the term“plasmapheresis” is often used interchangeably with therapeutic plasmaexchange and at other times it is used to denote a form of therapeuticplasma exchange where less plasma is removed than that removed intherapeutic plasma exchange. To avoid confusion with respect toterminology, the term plasmapheresis is used throughout, and, as stated,as used herein the term therapeutic plasma exchange is interchangeablewith the term plasmapheresis. However, in no way should the termplasmapheresis be deemed to be limiting on the scope of the disclosurefound herein which may be relevant to different types of apheresis basedon context.

A plasmapheresis therapy session may begin with the initial step ofwithdrawing whole blood from a blood vessel of a patient using anapheresis device. Apheresis devices are well known and are machinesconfigured to carry out procedures including plasmapheresis. Apheresisdevices may be configured to withdraw whole blood from an individualthrough an intravenous line, separate the whole blood into components,and return an infusion to the individual through an intravenous line.The infusion returned to the individual may include separate componentswhich may include blood that has had the plasma component removed fromit. In addition, an infusion given to the individual may include anexchange fluid as well. An apheresis device can be an ex vivo apheresissystem or machine comprising one or more centrifugal chambers. An exvivo apheresis system or machine can also comprise a return flowcontroller and one or more sensors for monitoring plasma or blooddensity. An apheresis device can also be configured to deliver ananticoagulant to the patient during the procedure. In some embodiments,the anticoagulant can be citrate dextrose. It should, however, beunderstood that any method or device for carrying out plasmapheresis issuitable for use with the methods and formulations described herein andthe description provided should not be deemed to limit the inventivemethods or formulations described herein which are suitable for use withany device or method for carrying out plasmapheresis.

An exchange fluid is typically administered with plasmapheresis whereinthe exchange fluid is administered to the individual receivingplasmapheresis intravascularly (i.e. through intravenous access; e.g.peripheral or central line) during the plasmapheresis treatment. Anexchange fluid may comprise any fluid that is suitable for use inintravenous fluid administration. For example, non-limiting examples offluids suitable for intravascular administration with the administrationof plasmapheresis as described herein are commonly referred to asisotonic fluids and include Normal Saline (i.e. a 0.9% saline solution)and Lactated Ringer's. An exchange fluid solution suitable for use inplasmapheresis as described herein may further include albumin such ashuman albumin. For example, an exchange fluid may comprise a normalsaline solution that includes 5% albumin by weight. Typically, a sourceof albumin is human derived albumin also referred to as human serumalbumin (HSA). As an example, an exchange fluid suitable for use withplasmapheresis may comprise a sterile liquid preparation comprising anamount of human-derived protein in the amount of 50 g per 1000 ml of thesterile liquid preparation wherein at least 96% of the human-derivedprotein is human serum albumin protein. Besides the human-derived serumalbumin protein, the remainder of the HSA preparation can comprise asaline solution and small amounts of potassium, N-acetyl-DL-tryptophan,caprylic acid, or a combination thereof. A 5% HSA preparation of anexchange fluid can be an FDA-approved 5% HSA preparation. Morespecifically, the 5% HSA preparation can be manufactured by anFDA-approved procedure such as the Cohn-Oncley cold ethanolfractionation procedure followed by ultra-filtration and pasteurization.Replacing plasma withdrawn from an individual receiving plasmapheresiswith 5% HSA is useful for regulating and stabilizing the volume ofcirculatory blood within the individual.

An exchange fluid may be mixed or combined in any suitable way with thecellular fraction of the whole blood that is withdrawn duringplasmapheresis, which remains after separation of plasma and which isreturned to the individual receiving plasmapheresis during theprocedure. An exchange fluid suitable for use with plasmapheresis mayalso include one or more therapeutics. For example, a therapeutic thatreduces inflammation may be provided with plasmapheresis by, forexample, mixing the therapeutic with an exchange fluid. An exchangefluid for plasmapheresis may also comprise or be mixed together one ormore blood products (i.e. not including the cellular fraction that isreturned) including but not limited to fresh frozen plasma andplatelets. It should be understood, that when mixed with an exchangefluid, therapeutics and blood products will to at least some degree bewithdrawn back from the individual receiving the plasmapheresis and assuch it may be beneficial to administer or infuse a therapeutic and/orblood product after completing the blood withdrawal from the individualso that the therapeutic and/or blood product will not be withdrawn fromthe individual during the plasmapheresis therapy.

As stated, in a typical plasmapheresis procedure, a volume of wholeblood is withdrawn from an individual and a portion of it is returned tothe individual along with an exchange fluid. Typically, the exchangefluid is returned to the individual simultaneously with the withdrawalof the whole blood which makes calculations related to theplasmapheresis process not entirely simple and straightforward.

Typically, as stated above, exchange fluid is infused to the individualreceiving plasmapheresis simultaneously with withdrawal of the wholeblood and removal of the plasma so that the exchange fluid isessentially reconstituting some of the plasma volume during theprocedure, and it is not, therefore, possible to remove the entireplasma volume during a typical plasmapheresis procedure because, to somedegree, plasma is being continuously replenished as it is being removed.As used herein, “plasma volume” refers to the entire volume of plasmawithin the whole blood of an individual, which can be calculated usingnumerous methods that are well known for calculating plasma volume. In atypical plasmapheresis procedure, a volume of plasma that is equal toabout 1 plasma volume is removed from the individual receivingplasmapheresis or 1 plasma volume up to 1.5 plasma volumes may beremoved in a typical plasmapheresis procedure. Typically, during aplasmapheresis administration as described herein, an amount of exchangefluid is returned to the patient that is essentially equal to the amountof plasma volume that is withdrawn from the patient so that the removedplasma volume (that is not returned to the patient) is “exchanged” withthe exchange fluid that is returned to the patient and replaces thevolume of plasma that is removed. While the exchange fluid in someembodiments may be more or less than the plasma volume removed, again,typically it is essentially equal in volume. For example, where oneplasma volume is removed from an individual in a method describedherein, a volume of exchange fluid equal (or essentially equal) to oneplasma volume is returned to the individual.

As explained, because exchange fluid is simultaneously infused to anindividual during a plasmapheresis procedure, removing anywhere from1-1.5 plasma volumes during a plasmapheresis procedure does nottypically mean that the plasma (and its contents) are completely removedeven though a volume equal to 1-1.5 plasma volumes is removed, becausethe removed volume includes exchange fluid that had been infused to theindividual and then removed as part of the total volume removed.According to Winters (Hematology Am Soc Hematol Educ Program, 2012;2012(1):7-12), in a typical plasmapheresis treatment where 1-1.5 ofplasma volume exchanged, approximately 60%-70% of substances present inthe plasma at the start of plasmapheresis will be removed which meansthat approximately 30-40% of substances present in the plasma at thestart of plasmapheresis will remain in the body of the individualreceiving plasmapheresis following a typical single plasmapheresistreatment.

Depending on the weight of the individual receiving plasmapheresis, thevolume of plasma removed can be between approximately 2 L to 4 L. When 2L to 4 L of plasma is removed during plasmapheresis, the volume of wholeblood that is withdrawn from the patient is necessarily greater than 2.0L to 4.0 L, which is to say that the withdrawn whole blood volumecontains the volume of plasma to be removed and therefore the wholeblood volume withdrawn is necessarily always larger than the volume ofplasma that is removed. It should be understood that plasma volume in anindividual is dependent on a number of factors including weight andgender and so 2 L to 4 L is used here only as a non-limiting example ofa range of plasma that might be removed in a plasmapheresis procedure.plasmapheresis, in some instances may involve removal of less than 2 Lof plasma or more than 4 L of plasma.

Blood can be withdrawn from a blood vessel of the patient including aperipheral blood vessel, a central blood vessel, or a combinationthereof. Since the blood flow from the patient into the apheresis devicehas to be steady and preferably faster than 50 mL/min, the site ofvascular access is typically a blood vessel capable of withstanding highnegative pressure without collapsing.

Moreover, a site of vascular access for receiving an exchange fluid or amixture that includes an exchange fluid and one or more other componentsis typically another blood vessel (or another peripheral or centralaccess point) capable of tolerating relatively high positive pressure.In some embodiments of the methods described herein, whole blood can bewithdrawn using a large-bore needle or cannula from a patient'speripheral vein such as the antecubital fossa, the basilica vein, or thecephalic vein. Additionally, if determined by a plasmapheresis providerto be an optimal vascular access point, whole blood can also bewithdrawn by cannulation of a radial artery of an individual receivingplasmapheresis. If determined by a plasmapheresis provider to be anoptimal vascular access point, whole blood can be withdrawn using anintravascular or implantable device such as a central venous catheter(CVC), an arteriovenous (AV) shunt, an AV fistulae, or a port-CVC. Forexample, whole blood can be withdrawn from an internal jugular vein, asubclavian vein, or a femoral vein or artery of the individual receivingplasmapheresis. Blood from an individual receiving plasmapheresis can bewithdrawn at a rate of approximately 90 ml/min. However it is alsosuitable, in the methods described herein, that blood from an individualreceiving plasmapheresis be withdrawn at a rate of between approximately90 ml/min and 135 ml/min. It should be understood that, under certainconditions, withdrawal at a rate of less than 90 ml/min or more than 135ml/min may be optimal for the individual receiving plasmapheresis. Aspreviously discussed, typically, a site of vascular access for receivinga fluid to be returned to an individual receiving plasmapheresis, whichmay comprise, for example, an exchange fluid, a cellular fraction, atherapeutic, or a blood product and mixtures thereof, is different froma site of vascular access for initial blood withdrawal. For example, acannula or catheter extending from or otherwise coupled to an apheresisdevice can be used to deliver a fluid to be returned to an individualreceiving plasmapheresis comprising a cellular fraction and an exchangefluid to a blood vessel in an arm, hand, neck, or chest of an individualreceiving plasmapheresis.

In some embodiments of the methods described herein, each plasmapheresistreatment session can last approximately 90 minutes to 2 hours. However,it should be understood that plasmapheresis session length can be variedbased on the objective of the plasmapheresis treatment and sessionsshorter than 90 minutes or longer than 2 hours are suitable with themethods and formulations described herein. In addition, a plasmapheresistreatment session duration can vary depending on certain factorsassociated with the individual receiving the plasmapheresis includingbut not limited to the weight of the individual receiving theplasmapheresis or the overall health of the individual.

In an exemplary method for administering plasmapheresis to anindividual, a plasmapheresis method can comprise one or a plurality ofplasmapheresis therapies. A plasmapheresis method can begin with thestep of identifying an individual in need of a plasmapheresis treatment.The treatment method can further comprise withdrawing whole blood from ablood vessel of the individual receiving plasmapheresis using anapheresis device or other technique for withdrawing blood. Theplasmapheresis method can further comprise separating the whole bloodwithdrawn into a cellular fraction and a plasma fraction using theapheresis device or other technique for separating blood components. Theplasmaphersis method can further comprise infusing back to theindividual receiving plasmapheresis an exchange fluid and the cellularfraction while removing the plasma fraction from the individual. Anamount of exchange fluid returned to the individual may be approximatelyequal to an amount of plasma that is removed. For example, if one plasmavolume is removed from an individual receiving plasmapheresis, incertain methods described herein, an amount of an exchange fluidreturned to the individual will also be approximately equal to oneplasma volume. Alternatively, an amount of exchange fluid returned mayalso exceed the amount of plasma volume removed. For example, if oneplasma volume is removed, more than one plasma volume may be infusedback to the individual receiving the plasmapheresis.

Factors involved in aging are located within the plasma of a patient andtherefore, using the innovative plasmapheresis methods described herein,the factors involved with aging are removed from the body of anindividual receiving plasmapheresis with removal of plasma from the bodyof the individual.

As explained, aging is likely a multifactorial process that currentlyhas not been fully elucidated, which has generally precluded developmentof effective therapies to treat and prevent the effects of aging.However, while the entire pathophysiology of aging may not yet becompletely clear, there are clear choke-points in the pathophysiology ofaging where the innovative methods described herein are effective. Forexample, it is understood that factors that affect aging includingcytokines and peptides associated with aging can all be found in plasma.As used herein “plasma content” describes all of the separate componentsof plasma, and we know that at least some of the plasma content hascomponents in it that affect aging (even if we don't know what they arespecifically within the plasma content). The presence of these agingfactor or factors within the plasma, makes the plasma a potential chokepoint of the aging process because these targeted aging-related factoror factors must necessarily travel through the vascular system withinplasma in order to get to the cells and/or tissue upon which they act,and, therefore, these factor or factors can be effectively collected andremoved from the vascular system choke point using the innovativemethods described herein. Even if it is not necessarily known whichfactor or factors found in blood are to be targeted by an age-relatedtherapy, removing all of the plasma content or as much as is possibleusing the innovative methods and formulations described herein alsonecessarily removes the factor or factors that affect aging from thebody and therefore diminishes or prevents the aging activity thatremoved factor or factors are associated with.

As already explained, a plasmapheresis session typically removes about60-70% of plasma content which means that about 30-40% of the plasmacontent in the body of the individual receiving a typical singleplasmapheresis treatment, including those factors associated with aging,remain within the blood of the individual receiving plasmapheresis aftera single plasmapheresis treatment.

In certain methods described herein, a plasmapheresis protocol removesmore plasma than is removed in a typical single plasmapheresis treatmentso that more than 60-70% of the plasma content of an individualreceiving plasmapheresis is removed from the body of the individual. Forexample, in certain methods described herein, a first plasmapheresistreatment may remove 60-70% of plasma content of an individual and theindividual then undergoes an additional plasmapheresis treatment withina window of time relative to the first plasmapheresis treatment so thatthe first plasmapheresis treatment together with the additionalplasmapheresis treatment achieves a cumulative removal of plasma greaterthan 60-70%. In these embodiments, the additional plasmapheresistreatment is administered before the plasma that is removed from theindividual who received the plasmapheresis is fully replenished in theblood of the individual who received the plasmapheresis. In theseembodiments, a plasmapheresis therapy method as described herein may becarried out and then a subsequent plasmapheresis therapy method asdescribed herein may be carried out a time that is within 72 hours ofthe initial plasmapheresis therapy. Similarly, a plasmapheresis therapymethod as described herein may be carried out and then an additionalplasmapheresis therapy method as described herein may be carried out atime that is within 48 hours of the initial plasmapheresis therapy. Forexample, a plasmapheresis therapy method as described herein may becarried out and then an additional plasmapheresis therapy method asdescribed herein may be carried out a time that is within 24 hours ofthe initial plasmapheresis therapy. Similarly, in these methods, anadditional plasmapheresis treatment may also be carried out within awindow of time where an additional plasmapheresis treatment in anindividual is within four days, within five days, within six days,within seven days, within 8 days, within 9 days, within 10 days, within11 days, within 12 days, within 13 days, or within 14 days of an initialplasmapheresis treatment in the individual.

Other approaches to achieving as much removal of plasma content as ispossible include withdrawing more than 1.5 times of the plasma volume ina single plasmapheresis session. For example, a method for performingplasmapheresis as described herein includes withdrawing 1.5 times theplasma volume of an individual receiving plasmapheresis in a singleplasmapheresis treatment. For example, a method for performingplasmapheresis as described herein includes withdrawing 1.6 times theplasma volume of an individual receiving plasmapheresis in a singleplasmapheresis treatment. For example, a method for performingplasmapheresis as described herein includes withdrawing 1.7 times theplasma volume of an individual receiving plasmapheresis in a singleplasmapheresis treatment. For example, a method for performingplasmapheresis as described herein includes withdrawing 1.8 times theplasma volume of an individual receiving plasmapheresis in a singleplasmapheresis treatment. For example, a method for performingplasmapheresis as described herein includes withdrawing 1.9 times theplasma volume of an individual receiving plasmapheresis in a singleplasmapheresis treatment. For example, a method for performingplasmapheresis as described herein includes withdrawing 2 times theplasma volume of an individual receiving plasmapheresis in a singleplasmapheresis treatment. Likewise, a method for performingplasmapheresis as described herein may include withdrawing 2 or moretimes the plasma volume of an individual receiving plasmapheresis in asingle plasmapheresis treatment.

In each case where a high percentage of plasma content is removed,clotting factors and/or platelets may be infused back to the individualreceiving the plasmapheresis to replenish the clotting factors andplatelets removed with the plasma content to address any elevatedbleeding risk post the plasmapheresis treatment due to removal of theclotting factors and platelets.

A “plasmapheresis regimen” as used herein includes within its scope anyapplication of plasmapheresis as described herein for the treatment orprevention of aging and/or promotion of longevity. A plasmapheresisregimen may include one or more treatments carried out over a particulartime period and may in certain implementations include therapeuticsprovided together with plasmapheresis.

In the innovative methods described herein, plasmapheresis may be usedin a plasmapheresis regimen in order to remove a relatively large amountof plasma content from an individual receiving plasmapheresis. Incertain plasmapheresis regimens, plasmapheresis may be administered on atwice a week schedule to achieve a cumulative effect with respect toplasma content removal as described herein with the time period betweena first and a second plasmapheresis treatment being delivered to anindividual within 24 hours of each other (including within the sameday), within 36 hours of each other, within 48 hours of each other,within 60 hours of each other, within 72 hours of each other, within 84hours of each other, within 96 hours of each other, within 108 hours ofeach other, within 120 hours of each other, within 134 hours of eachother, within 156 hours of each other, or within 168 hours of eachother. An additional plasmapheresis treatment may also be carried outwithin a window of time where an additional plasmapheresis treatment inan individual is within four days, within five days, within six days,within seven days, within eight days, within nine days, within ten days,within eleven days, within twelve days, within thirteen days, or withinfourteen days of an initial plasmapheresis treatment in the individual.

A plasmapheresis regimen as described that is carried out twice a monthmay be performed every week in a month for a total of eightplasmapheresis treatments per month, every other week for a total offour plasmapheresis treatments per month, or once a month for a total oftwo treatments in a month. A plasmapheresis regimen as described hereinwith twice weekly treatments may be carried out for 6 months in total oftwice weekly treatments. Within that 6 month period the twice weeklytreatments may all be carried out with the same amount of time betweeneach of the two weekly treatments or the amount of time may be variedbetween the two weekly treatments.

A plasmapheresis regimen as described herein may comprise performance ofplasmapheresis once per week. For example, a plasmapheresis regimen asdescribed that is carried out once a month may be performed once a weekevery week in a month for a total of four plasmapheresis treatments permonth, every other week for a total of two plasmapheresis treatments permonth, or once a month for a total of one treatment in a month. Anyplasmapheresis regimen as described herein with once weekly treatmentsmay be carried out for 6 months in total.

In certain plasmapheresis treatments described herein, any subsequentplasmapheresis treatment session can be undertaken only after 24 dayshave passed since the last plasmapheresis treatment session. In theseand other embodiments, the entire treatment method can cease after 125days have passed since the first plasmapheresis treatment session. Thetreatment method can also comprise continuing the treatment method byrepeating the method when at least 24 days have passed since the lasttreatment session. In these embodiments, no plasmapheresis treatmentsessions should occur during this intervening waiting period. Forexample, the various plasmapheresis treatment sessions can be separatedby 24 days, 25 days, 26 days, 27 days, 28 days, 29 days, or anycombination thereof. The various plasmapheresis treatment sessions canbe separated by the same number of days or be separated by a differingnumber of days. As a more specific example, the first plasmapheresistreatment session can be separated by the second plasmapheresistreatment session by 24 days and the second plasmapheresis treatmentsession can be separated by the third plasmapheresis treatment sessionby 25 days or 26 days. In other example methods, the firstplasmapheresis treatment session can be separated by the secondplasmapheresis treatment session by 26 days and the secondplasmapheresis treatment session can be separated by the thirdplasmapheresis treatment session by 25 days or 24 days. In someembodiments, the treatment method can cease entirely when 125 days havepassed since the first treatment session. In other embodiments, thetreatment method can cease when at least six plasmapheresis treatmentsessions have been undertaken, regardless of the number of days passedsince the first plasmapheresis treatment session. In additionalembodiments, the treatment method can cease when at least seven or atleast eight plasmapheresis treatment sessions have been undertaken,regardless of the number of days passed since the first plasmapheresistreatment session. In another treatment method for performingplasmapheresis, the treatment method can comprise six (6) plasmapheresistreatment sessions in total. The first plasmapheresis treatment sessioncan occur on the first day (day 1) of the treatment, the secondplasmapheresis treatment session can occur on the twenty-fifth day (day25) of the treatment method, the third plasmapheresis treatment sessioncan occur on the fiftieth day (day 50) of the treatment method, thefourth plasmapheresis treatment session can occur on the seventy-fifthday (day 75) of the treatment method, the fifth plasmapheresis treatmentsession can occur on the one-hundredth day (day 100) of the treatmentmethod, and the sixth plasmapheresis treatment session can occur on theone-hundred and twenty-fifth day (day 125) of the treatment method. Inthis embodiment, no plasmapheresis treatment sessions occur during theintervening periods between the aforementioned treatment sessions. Inother embodiments, a plasmapheresis treatment regimen may extend as longas needed and include as many separate sessions/treatments as needed toachieve one or more measurable effects.

With respect to the use of plasmapheresis in the treatment of aging andaging related conditions, an anti-inflammatory or otherimmune-modulating therapeutic may be used together with plasmapheresistreatment in a synergistic way. More specifically, an anti-inflammatoryor immune-modulating therapeutic may synergistically modulate, reduce,or eliminate the effect of inflammatory and immune cells and factors(e.g. cytokines) that remain in the blood following plasmapheresistreatment and thereby further reducing the effect of these cells andfactors. In certain of the methods described herein, a therapeutic isadministered to target an aspect of aging found within blood of anindividual receiving the plasmapheresis treatment. For example,inflammation is a known part of the aging process and is sometimesreferred to as “inflammaging” wherein effects of aging correlate with anelevated level of inflammatory agents within the blood. As such, ananti-inflammatory delivered together with plasmapheresis can act toreduce inflammatory activity and as such disrupt the effect ofinflammation in age related conditions. With plasmapheresis inparticular, there's a strong synergy in that using the methods describedherein most of the non-cellular inflammatory factors found within theplasma content are removed from the body of the individual receiving theplasmapheresis treatment and any remaining inflammatory cells may befurther affected with therapeutics. As such, in the example provided,with a single typical plasmapheresis about 30-40% of the inflammatoryfactors within the plasma content will remain after the single typicalplasmapheresis treatment and if an anti-inflammatory or otherimmune-modulator is administered with or soon after plasmapheresis isperformed, the anti-inflammatory or other immune-modulator can actsynergistically with the removal of the inflammatory factors to furtherattenuate the effect of the inflammatory cells and factors remainingafter the plasmapheresis treatment. Applying a therapy in this targetedway creates a cumulative inhibition effect that increases the benefit tothe individual receiving the plasmapheresis.

A plasmapheresis treatment as described herein can further comprisedelivering a therapeutically-effective dosage of intravenousimmunoglobulin (IVIG) to a blood vessel of the individual receiving theplasmapheresis treatment. In certain plasmapheresis treatments asdescribed herein, IVIG is delivered to an individual receivingplasmapheresis after returning the cellular fraction and the exchangefluid to the individual receiving the plasmapheresis. This is to saythat a therapeutically-effective dosage of IVIG may be provided to anindividual separately from infusion of the cellular fraction and theexchange fluid to the individual receiving the plasmapheresis. Forexample, IVIG may be infused in the individual receiving plasmapheresisafter a plasmapheresis treatment is completed so that any infused IVIGwill not be removed in the plasmapheresis process. It is also possibleto administer IVIG concurrently with the administration ofplasmapheresis.

The therapeutically-effective dosage of IVIG can be approximately 2.0 gof IVIG per kg of bodyweight of the individual receiving plasmapheresis.The IVIG delivered can contain certain antibodies and cytokines whichcan have a positive effect on the immune system of the individualreceiving plasmapheresis and may contribute to establishing an optimalsystemic environment for cell growth. As stated above, it is suitablefor administration of a therapeutic such as IVIG mixed together with anexchange fluid and/or a cellular fraction during a portion of theplasmapheresis procedure when blood is being withdrawn or it is alsosuitable to administer IVIG to the individual as part of a method foradministering plasmapheresis as described herein when blood is no longerbeing withdrawn from the individual receiving plasmapheresis. Thetherapeutically-effective dosage of IVIG can be any FDA-approved IVIG orimmune globulin intravenous (IGIV) infusion or preparation. For example,the IVIG can comprise primarily of gamma globulins.

The steps described do not require the particular order shown to achievethe desired result. Moreover, certain steps or processes may be omittedor occur in parallel in order to achieve the desired result.

Each of the plasmapheresis treatment sessions can comprise the steps ofwithdrawing whole blood from a blood vessel of a patient using anapheresis device, separating the whole blood withdrawn into a cellularfraction and a plasma fraction using the apheresis device, admixing thecellular fraction with an exchange fluid comprising albumin derived fromhuman plasma, returning the mixture comprising the cellular fraction andthe exchange fluid to the blood vessel of the patient using an apheresisdevice, and delivering a therapeutically-effective dosage of intravenousimmunoglobulin (IVIG) to the blood vessel of the individual receivingplasmapheresis after returning the mixture comprising the cellularfraction and the exchange fluid to a blood vessel of the individual.

A biological age or any other measurable feature or marker of theefficacy of plasmapheresis on an individual may be measured using thenumerous markers and assays described herein. In particular, abiological age, physiological measurement (e.g. strength, walking, orbalance), mental assessment (e.g. an emotional wellness survey), ormarker in the blood of the individual (e.g. cell surface marker) that ismeasured using any of the analysis techniques, markers, and assaysdescribed herein may be used in conjunction with a plasmapheresisregimen as described herein wherein a biological age, physiologicalmeasurement, mental assessment, or marker that is identified,quantified, or determined to be present is used to affect a modificationin a plasmapheresis regimen.

For example, a biological age, physiological measurement, mentalassessment, or marker in the blood of an individual may be measured,using any of the markers and/or assays described herein, before andafter a plasmapheresis regimen as described herein (including one ormore plasmapheresis treatments over a period of time) and theplasmapheresis regimen may be extended if a decrease (or increase if anincrease and not a decrease is beneficial with respect to the item thatis measured) in the biological age, physiological measurement, mentalassessment, or marker in the blood of the individual (as measured beforethe regimen is administered) is not achieved. The plasmapheresis regimencan then be continued until the decrease (or increase if an increase andnot a decrease is beneficial with respect to the item that is measured)in the biological age, physiological measurement, mental assessment, ormarker that is sought is achieved.

Similarly, biological age, a physiological measurement, a mentalassessment, or a marker found in the blood of an individual can shapethe plasmapheresis regimen itself, wherein, for example, a biologicalage, a physiological measurement, a mental assessment, or a marker inthe blood of an individual is measured before and after a singleplasmapheresis therapy and subsequent plasmapheresis therapies areadministered until the biological age is decreased below a level that issought.

In an exemplary method, a plasmapheresis method can begin with the stepof identifying an individual in need of a plasmapheresis therapy. Ablood sample is taken from the individual, either before initiation ofthe plasmapheresis, or from the whole blood that is withdrawn and usedto determine a biological age of the individual using the markers and/orassays described herein. In some embodiments, physiological and mentalassessment is carried to assess one or more of the individual'sstrength, walking, balance, or mental status (wherein exemplarytechniques for obtaining these measurements are described herein such asin the section below describing the pilot study and results). Theplasmapheresis method can further comprise withdrawing whole blood froma blood vessel of the individual receiving plasmapheresis using anapheresis device or other technique for withdrawing blood. Theplasmapheresis method can further comprise separating the whole bloodwithdrawn into a cellular fraction and a plasma fraction using theapheresis device or other technique for separating blood components. Thetreatment method can further comprise infusing back to the individualreceiving plasmapheresis an exchange fluid and the cellular fractionwhile removing the plasma fraction from the individual. An amount ofexchange fluid returned to the individual may be approximately equal toan amount of plasma that is removed. For example, if one plasma volumeis removed from an individual receiving plasmapheresis, in certainmethods described herein, an amount of an exchange fluid returned to theindividual will also be approximately equal to one plasma volume.Alternatively, an amount of exchange fluid returned may also exceed theamount of plasma volume removed. For example, if one plasma volume isremoved, more than one plasma volume may be infused back to theindividual receiving the plasmapheresis. Once the plasmapheresis therapyis completed, a second sample is obtained and a second biological age isdetermined using the markers and/or assays described herein. Should oneor more of the biological age, physiological measurement, mentalassessment, or marker found in the blood of the individual be determinedafter the plasmapheresis therapy not to have been lowered or raisedsufficiently (as compared to the biological age, physiologicalmeasurement, mental assessment, or the marker determined before thestart of the plasmapheresis therapy), additional plasmapheresis therapyis administered with repeated sampling and measuring of biological age,physiological measurement, mental assessment, or the marker with theplasmapheresis continuing until the biological age, physiologicalmeasurement, mental assessment, or the marker is decreased by an amountthat is sought.

Also described herein is a method for treating aging by carrying out ablood dilution. As described herein, plasma content can be removed andexchange fluid added. The removal of plasma content (even with partialreplacement) together with the addition of exchange fluid typicallylowers the concentrations of one or more constituents of plasma in anindividual receiving plasmapheresis treatment. Essentially, solute (theplasma content) is decreased while the solvent (the liquid portion ofplasma replaced by the exchange fluid) remains the same. In this way, inaddition to removing plasma content components, the methods describedherein dilute one or more components of plasma content that remain inthe blood of the individual receiving plasmapheresis following aplasmapheresis treatment.

Plasma content dilution can achieve synergistic affects to removal ofplasma content alone in that, at least, dilution causes the plasmacomposition of an individual receiving plasmapheresis to resemble aplasma composition of a biologically younger individual. This is to saythat factors found in plasma that are associated with aging are foundempirically to have a lower concentration in younger individuals thanolder individuals. As in a younger individual where the aging-associatedfactors found in plasma are less active, the decrease in concentrationof the plasma content in individuals receiving plasmapheresis asdescribed herein lowers the relative activity further of the factorsassociated with aging in those individuals receiving the plasmapheresistherapy as described herein.

(i) Modeling Blood Dilution

A number of models may be used to calculate an amount of dilution thatis achieved with a plasmapheresis treatment as described herein. Forexample, Reverberi and Reverberi (Blood Transfus, 2007; 5(3):164-174)provide formulas for modeling residual plasma analyte concentrationsfrom which Formulas (I), (Ia), and (Ib) are derived:

$\begin{matrix}{{{Residual}{Solute}{Concentration}} = {100\%*e^{- \frac{\nu_{P}*n}{\nu_{b}}}}} & (I)\end{matrix}$

wherein v_(p) is plasma volume, n is number of plasma volume unitsexchanged, and v_(b) is total blood volume (i.e. whole blood volume).

As an illustrative example of Formula (I), using a plasma volume of 3 L,1 plasma volume to be exchanged, and a total volume of 5 L of wholeblood would be expected to result in a residual plasma contentconcentration of approximately 54% of the original concentrationfollowing a single plasmapheresis treatment. For the same plasma volumeand total volume of blood but a plasma exchange of 1.5 plasma volumes,the residual solute concentration would be approximately 40%.

Typically, plasma analytes exhibit an instantaneous concentration dropsfollowing blood exchange, followed by reconcentration along alogarithmic-like trends to pre-blood dilution levels. Formula (I) can bemodified to Formula (Ia) to estimate plasma analyte levels n_(d) daysfollowing plasma exchange, assuming a half-time t_(1/2) for the analytesto return to pre-blood dilution levels:

$\begin{matrix}{{{Residual}{Solute}{Concentration}} = {100\%*{( {1 - {\begin{pmatrix}1 \\ - \\2\end{pmatrix}^{\frac{n_{d}}{t_{\frac{1}{2}}}}e^{- \frac{\nu_{p}*n}{\nu_{b}}}}} ).}}} & ({Ia})\end{matrix}$

Noting that not all plasma analytes reestablish homeostasis at equalrates, as typical plasma analytes return to pre-blood dilution levelsafter about 10 days, t_(1/2) of 3 to 4 days is often a suitableestimate. Once again using 3 L, 1 plasma volume, and 5 L of whole bloodwith a half-life of 3 days and measured 3 days after the initialplasmapheresis treatment, the residual solute concentration increasesfrom approximately 54% following the plasmapheresis treatment toapproximately 72% three days (i.e. 72 hours) later.

Winters (cited above) provides the following simplified formula:Y/Y0=e^(−x), where Y is the final concentration of a substance, Y0 isthe initial concentration, and X is the number of times the patient'splasma volume is exchanged. Continuing with the approximately 72%residual concentration calculated above using Formula (Ia), ifplasmapheresis is administered once again 72 hours after the initialplasmapheresis, Y0=72%, x=1, and Y=26.5% residual plasma concentrationfollowing the second plasmapheresis treatment.

Formula (Ia) can further be used to determine the degree of plasmaanalyte clearance following multiple rounds of plasma exchanges bysetting n as above. For such an application, Formula (Ia) can beestimated as a sum, expressed as Formula (Ib), where each plasmaexchange event, i, is attenuated by analyte regeneration over n_(di)days following the plasma exchange event:

$\begin{matrix}{{{Residual}{Solute}{Concentration}} = {100\%*{( {1 - {\sum_{i}{\begin{pmatrix}1 \\ - \\2\end{pmatrix}^{\frac{n_{di}}{t_{\frac{1}{2}}}}e^{- \frac{\nu_{p}}{\nu_{b}}}}}} ).}}} & ({Ib})\end{matrix}$

This equation does not account for diminished plasma exchange efficacyover multiple cycles, reflecting exchange of partially diluted plasma.However, in cases where t_(1/2) is approximately equal to or less thanthe time between plasma exchange events, and for therapies with limitednumbers of plasma exchanges, discrepancies from this estimation aretypically small.

Formulas (I), (Ia), (Ib), and the formula provided by Winters allprovide excellent and useful approximations of dilution levels which aresuitable for use with the methods described herein. It is also notedthat if higher accuracy in calculating plasma content dilution should beachieved, the following factors and issues should also be considered:First, many plasma separation methods only partially separate plasmafrom cellular components. While multiple rounds of separation canincrease efficiencies, single iterations of centrifuge-based plasmaseparation and membrane-based plasma separation typically achieve 80%and 30% plasma separation efficiencies (Williams and Balogun. Clin J AmSoc Nephrol, 2014; 9(1): 181-190). For a 500 mL blood draw, which willtypically contain about 55% (275 mL) plasma these efficiencies translateto 220 mL plasma removed by centrifugation and 82.5 mL plasma removed byfiltration. The effect of incomplete plasma separation from cellularcomponents is a concomitant decrease in plasma exchange efficiency. Ifplasma is centrifugally separated from blood with 80% efficiency, thenplasma exchange will typically be 80% efficient given a volume of bloodseparated and exchanged. Second, many plasma components activelyexchange into spaces outside of the vasculature. While a typical adultmale human has about 5 liters of blood, the majority of fluids, andanalogously, analytes, are contained in interstitial and intracellularspaces, which account for about 10.5 and 28 liters of fluid,respectively. As used herein, intracellular spaces can include allvolumes contained within cell membranes of an organism, including allfluids within these spaces, while interstitial spaces can denote spacessurrounding tissues. Many plasma analytes actively equilibrate betweenthe blood and these spaces, such that fractions of their totalpopulations are contained within the blood at any given time. Removal ofsuch species by plasma exchange can be attenuated by their partitioningoutside of the blood. For example, only about 60% to 70% of IgG1immunoglobulins are present in the blood at any given time, such thatplasma exchange can only target 60% to 70% of the IgG1 population.Furthermore, disruption of homeostasis, for example through blooddilution, can affect osmotic gradients which draw species into bloodfrom extravascular spaces, thereby hastening return to pre-treatmentblood analyte levels. Third, plasma analytes regenerate at a range ofrates. Following a blood composition altering event, such as blooddilution, blood analyte concentrations tend to return to their original,resting levels. Although blood dilution can alter resting levels ofindividual blood analytes (for example lowered blood triglyceride levelsas outlined in Dehal and Adashek. Case Rep Med, 2018; 2018: 4017573),diluted species tend to increase in concentration while concentratedspecies (e.g., albumin provided from a high concentration exchangefluid) tend to decrease in concentration to reestablish pre-dilutionlevels. While some species (in particular many cytokines) exhibitcomplex re-equilibration patterns, many follow simple exponential growthor decay curves. However, the rates of these processes can varysignificantly between species. For example, IgG immunoglobulins oftenreturn to pre-blood dilution levels in about 4 days (Harris et al.Journal of Scleroderma and Related Disorders, 2018; 3(2):132-152), whilelow-density lipoproteins can take more than 2 weeks to return to restinglevels (McGowan. Journal of Clinical Lipidology, 2013; 7(3):S21-S26).

May et al. (Am J Clin Pathol, 1989; 91(6):688-94) presents a model whichaccounts for extravascular compartmentalization and speciesregeneration. This model, adapted as Formula (II) below, captures ratesof plasma analyte loss through plasma exchange and clearance:

$\begin{matrix}{{A_{t} = {{A_{0}*e^{\lbrack{- \frac{{({r + k_{1}})}t}{V_{p}}}\rbrack}} + \frac{V_{p}{k_{2}\lbrack {1 - e^{\lbrack{- \frac{{({r + k_{1}})}t}{V_{p}}}\rbrack}} \rbrack}}{r + k_{1}}}},} & ({II})\end{matrix}$

wherein A_(t) denotes a plasma analyte concentration at time t, A₀denotes the initial concentration of the analyte, k₁ denotes a rate ofclearance of the analyte (e.g., through degradation, biliary clearance,etc.), and k₂ denotes a rate at which the analyte is released fromextracellular tissues. The May et al. model does not account forregeneration of the analyte, and further assumes plasma analytedistribution according to a two-compartment system (intravascular andextravascular). Accordingly, the model may not be appropriate foranalytes dynamically exchanged across multiple compartments (e.g.,antibodies with appreciable endosomal and interstitial concentrations),or analytes which are rapidly produced following dilution. However, thismodel can be corrected for inefficient plasma removal during exchange,as outlined above, through correction to clearance rate (k₁).

Human Pilot Study Results

An IRB approved pilot study was carried out to assess the effect ofplasmapheresis on aging in human participants. The study included eighthuman participants over the age of 50 (in chronological years) who eachreceived six plasmapheresis treatments over a three month period. Threeof the eight treated participants were female and five were male. Inaddition, a group of three participants each received three shamplasmapheresis treatments over three months and served as controls forthe eight participants who each received six plasmapheresis treatments.All three participants who received sham treatments were male.

The administration of the plasmapheresis treatments was overseen by alicensed physician using Spectra Optia plasma cell separatorsmanufactured by Terumo BCT, Inc., and FDA approved for that purpose.Plasmapheresis was administered in a clinical setting to the eightparticipants receiving the plasmapheresis treatment so that during eachof the six plasmapheresis treatments at least one plasma volume wasexchanged with an exchange fluid comprising 5% albumin over 2-3 hours.The three sham patients, used as controls, were given the appearance ofreceiving plasmapheresis in a clinical setting but did not actuallyreceive plasmapheresis.

Data that was collected and analyzed included macro data and micro data.macro data included measures of strength, balance, walking, and mentaland emotional wellbeing. micro data included blood studies that measuredand quantified cell surface markers associated with aging.

Macro Data

For the macro data, each participant had data collected before each oftheir six plasmapheresis treatments. This included measures of strength,balance, walking, and mental and emotional wellbeing. For strength, handgrip strength was measured using a grip strength measuring device. Forbalance, the participants were asked to stand and balance on one leg andthe total time they were able to stand on the one leg was recorded for amaximum time of 120 seconds. For walking, the participants stood from aseated position and walked a set distance and the total time it took theparticipant to stand and walk the distance was recorded. For mental andemotional wellbeing the participants answered questions from the SF-12survey, a recognized large-data-developed quality-of-life questionnaireconsisting of questions that measure emotional and mental health.

FIG. 1 shows multiple graphs of macro data for a first participant(“PT1”) in the pilot study who is female and who received six treatmentsof plasmapheresis. PT1 had macro data collected before each one of hersix plasmapheresis treatments. The exchange fluid of PT1 included 2grams of IVIG for each plasmapheresis treatment that PT1 received. Thegraphs of FIG. 1 are titled to indicate which macro data they present.For each of the five graphs shown in FIG. 1 , the x-axis has numbers 1through 6 indicating a time before each of the six plasmapheresistreatments when data was collected. For the graph titled “PT1 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, PT1'smeasured hand strength increased overall over the course of the sixtreatments. For the graph titled “PT1 Up&Go” the y-axis is the time inseconds that it took PT1 to stand from a seated position and walk afixed distance that PT1 walked each of the six times that themeasurement was taken (each participant tested also walked the samedistance). As can be seen, PT1's time to walk the distance decreasedbetween the first measurement and the last measurement indicating animprovement. For the graph titled “PT1 Balance” the y-axis is the timein seconds that PT1 was able to stand with one leg raised and where themaximum time measured was 120 seconds. As can be seen, PT1 was able tostand the maximum measured time throughout the duration of the study.For the graph titled “PT1 SF-12 Physical Health” the y-axis representsan SF-12 survey score for subjective physical health. As can be seen,PT1 showed an overall improvement in scores over the course of the sixtreatments. For the graph titled “PT1 SF-12 Emotional Health” the y-axisrepresents an SF-12 survey score for subjective emotional health. As canbe seen, PT1 showed an overall improvement in scores over the course ofthe six treatments.

FIG. 2 shows multiple graphs of macro data for a second participant(“PT2”) in the pilot study who is male and who received six treatmentsof plasmapheresis. PT2 had macro data collected before each one of hissix plasmapheresis treatments. The graphs of FIG. 2 are titled toindicate which macro data they present. For each of the five graphsshown in FIG. 2 , the x-axis has numbers 1 through 6 indicating a timebefore each of the six plasmapheresis treatments when data wascollected. For the graph titled “PT2 Hand Grip”, the y-axis shows themeasurements of grip strength in units of Kg measured with a standardgrip strength device. As can be seen, PT2's measured hand strengthincreased overall over the course of the six treatments. For the graphtitled “PT2 Up&Go” the y-axis is the time in seconds that it took PT2 tostand from a seated position and walk a fixed distance that PT2 walkedeach of the six times that the measurement was taken (each participanttested also walked the same distance). As can be seen, PT2's time towalk the distance decreased between the first measurement and the lastmeasurement indicating an improvement. For the graph titled “PT2Balance” the y-axis is the time in seconds that PT2 was able to standwith one leg raised and where the maximum time measured was 120 seconds.As can be seen, PT2 was able to stand the maximum measured timethroughout the duration of the study. For the graph titled “PT2 SF-12Physical Health” the y-axis represents an SF-12 survey score forsubjective physical health. As can be seen, PT2 showed an overallimprovement in scores over the course of the six treatments. For thegraph titled “PT2 SF-12 Emotional Health” the y-axis represents an SF-12survey score for subjective emotional health. As can be seen, PT2 showedan overall improvement in scores over the course of the six treatments.

FIG. 3 shows multiple graphs of macro data for a third participant(“PT3”) in the pilot study who is female and who received six treatmentsof plasmapheresis. PT3 had macro data collected before each one of hersix plasmapheresis treatments. The exchange fluid of PT3 included 2grams of IVIG for each plasmapheresis treatment that PT3 received. Thegraphs of FIG. 3 are titled to indicate which macro data they present.For each of the five graphs shown in FIG. 3 , the x-axis has numbers 1through 6 indicating a time before each of the six plasmapheresistreatments when data was collected. For the graph titled “PT3 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, PT3'smeasured hand strength measurements decreased from an initially highmeasurement and then rose and appeared to plateau at a lower value thanthe initial strength measurement through the course of the sixtreatments. For the graph titled “PT3 Up&Go” the y-axis is the time inseconds that it took PT3 to stand from a seated position and walk afixed distance that PT3 walked each of the six times that themeasurement was taken (each participant tested also walked the samedistance). As can be seen, PT3's time to walk the distance decreasedbetween the first measurement and the last measurement indicating animprovement. For the graph titled “PT3 Balance” the y-axis is the timein seconds that PT3 was able to stand with one leg raised and where themaximum time measured was 120 seconds. As can be seen, PT3's ability tobalance on one leg increased overall through the course of the sixtreatments. For the graph titled “PT3 SF-12 Physical Health” the y-axisrepresents an SF-12 survey score for subjective physical health. As canbe seen, PT3 showed an overall improvement in scores over the course ofthe six treatments. For the graph titled “PT3 SF-12 Emotional Health”the y-axis represents an SF-12 survey score for subjective emotionalhealth. As can be seen, PT3 showed an overall improvement in scores overthe course of the six treatments. FIG. 4 shows multiple graphs of macrodata for a fourth participant (“PT4”) in the pilot study who is femaleand who received six treatments of plasmapheresis. PT4 had macro datacollected before each one of her six plasmapheresis treatments. Thegraphs of FIG. 4 are titled to indicate which macro data they present.For each of the five graphs shown in FIG. 4 , the x-axis has numbers 1through 6 indicating a time before each of the six plasmapheresistreatments when data was collected. For the graph titled “PT4 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, PT4'smeasured hand strength increased overall over the course of the sixtreatments. For the graph titled “PT4 Up&Go” the y-axis is the time inseconds that it took PT4 to stand from a seated position and walk afixed distance that PT4 walked each of the six times that themeasurement was taken (each participant tested also walked the samedistance). As can be seen, PT4's time to walk the distance decreasedbetween the first measurement and the last measurement indicating animprovement. For the graph titled “PT4 Balance” the y-axis is the timein seconds that PT4 was able to stand with one leg raised and where themaximum time measured was 120 seconds. As can be seen, PT4's ability tobalance on one leg increased overall through the course of the sixtreatments. For the graph titled “PT4 SF-12 Physical Health” the y-axisrepresents an SF-12 survey score for subjective physical health. As canbe seen, PT4 showed an overall improvement in scores over the course ofthe six treatments. For the graph titled “PT4 SF-12 Emotional Health”the y-axis represents an SF-12 survey score for subjective emotionalhealth. As can be seen, PT4 showed an overall improvement in scores overthe course of the six treatments.

FIG. 5 shows multiple graphs of macro data for a fifth participant(“PT5”) in the pilot study who is male and who received six treatmentsof plasmapheresis. PT5 had macro data collected before each one of hissix plasmapheresis treatments. The graphs of FIG. 5 are titled toindicate which macro data they present. For each of the five graphsshown in FIG. 5 , the x-axis has numbers 1 through 6 indicating a timebefore each of the six plasmapheresis treatments when data wascollected. For the graph titled “PT5 Hand Grip”, the y-axis shows themeasurements of grip strength in units of Kg measured with a standardgrip strength device. As can be seen, PT5's measured hand strengthincreased overall over the course of the six treatments. For the graphtitled “PT5 Up&Go” the y-axis is the time in seconds that it took PT5 tostand from a seated position and walk a fixed distance that PT5 walkedeach of the six times that the measurement was taken (each participanttested also walked the same distance). As can be seen, PT's time to walkthe distance decreased between the first measurement and the lastmeasurement indicating an improvement. For the graph titled “PT5Balance” the y-axis is the time in seconds that PT5 was able to standwith one leg raised and where the maximum time measured was 120 seconds.As can be seen, PT5's ability to balance on one leg increased overallthrough the course of the six treatments. For the graph titled “PT5SF-12 Physical Health” the y-axis represents an SF-12 survey score forsubjective physical health. As can be seen, PT5 showed an overallimprovement in scores initially over the course of the six treatmentsthat then decreased. For the graph titled “PT5 SF-12 Emotional Health”the y-axis represents an SF-12 survey score for subjective emotionalhealth. As can be seen, PT5 showed an overall improvement in scores overthe course of the six treatments.

FIG. 6 shows multiple graphs of macro data for a fourth participant(“PT6”) in the pilot study who is male and who received six treatmentsof plasmapheresis. PT6 had macro data collected before each one of hissix plasmapheresis treatments. The exchange fluid of PT6 included 2grams of IVIG for each plasmapheresis treatment that PT6 received. Thegraphs of FIG. 6 are titled to indicate which macro data they present.For each of the five graphs shown in FIG. 6 , the x-axis has numbers 1through 6 indicating a time before each of the six plasmapheresistreatments when data was collected. For the graph titled “PT6 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, PT6'smeasured hand strength decreased initially and then increased so thatoverall over the course of the six treatments there was no measuredoverall change. For the graph titled “PT6 Up&Go” the y-axis is the timein seconds that it took PT6 to stand from a seated position and walk afixed distance that PT6 walked each of the six times that themeasurement was taken (each participant tested also walked the samedistance). As can be seen, PT6's time to walk the distance decreasedbetween the first measurement and the last measurement indicating animprovement. For the graph titled “PT6 Balance” the y-axis is the timein seconds that PT6 was able to stand with one leg raised and where themaximum time measured was 120 seconds. As can be seen, PT6's ability tobalance on one leg increased overall through the course of the sixtreatments. For the graph titled “PT6 SF-12 Physical Health” the y-axisrepresents an SF-12 survey score for subjective physical health. As canbe seen, PT6 showed an overall improvement in scores over the course ofthe six treatments. For the graph titled “PT6 SF-12 Emotional Health”the y-axis represents an SF-12 survey score for subjective emotionalhealth. As can be seen, PT6 showed an overall slight decrease in scoresover the course of the six treatments.

FIG. 7 shows multiple graphs of macro data for a fourth participant(“PT7”) in the pilot study who is male and who received six treatmentsof plasmapheresis. PT7 had macro data collected before each one of hissix plasmapheresis treatments. The graphs of FIG. 7 are titled toindicate which macro data they present. For each of the five graphsshown in FIG. 7 , the x-axis has numbers 1 through 6 indicating a timebefore each of the six plasmapheresis treatments when data wascollected. For the graph titled “PT7 Hand Grip”, the y-axis shows themeasurements of grip strength in units of Kg measured with a standardgrip strength device. As can be seen, PT7's measured hand strengthdecreased initially and then increased so that overall over the courseof the six treatments there a slight decrease in overall measuredstrength. For the graph titled “PT7 Up&Go” the y-axis is the time inseconds that it took PT7 to stand from a seated position and walk afixed distance that PT7 walked each of the six times that themeasurement was taken (each participant tested also walked the samedistance). As can be seen, PT7's time to walk the distance decreasedbetween the first measurement and the last measurement indicating animprovement. For the graph titled “PT7 Balance” the y-axis is the timein seconds that PT7 was able to stand with one leg raised and where themaximum time measured was 120 seconds. As can be seen, PT7 was able tostand the maximum measured time throughout the duration of the study.For the graph titled “PT7 SF-12 Physical Health” the y-axis representsan SF-12 survey score for subjective physical health. As can be seen,PT7 showed an overall improvement in scores initially over the course ofthe six treatments that then decreased. For the graph titled “PT7 SF-12Emotional Health” the y-axis represents an SF-12 survey score forsubjective emotional health. As can be seen, PT7 showed an overallslight decrease in scores over the course of the six treatments.

FIG. 8 shows multiple graphs of macro data for an eight participant(“PT8”) in the pilot study who is male and who received six treatmentsof plasmapheresis. PT8 had macro data collected before each one of hissix plasmapheresis treatments. The exchange fluid of PT8 included 2grams of IVIG for each plasmapheresis treatment that PT8 received. Thegraphs of FIG. 8 are titled to indicate which macro data they present.For each of the five graphs shown in FIG. 8 , the x-axis has numbers 1through 6 indicating a time before each of the six plasmapheresistreatments when data was collected. For the graph titled “PT8 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, PT8'smeasured hand strength decreased initially and then increased slightlybut overall decreased over the course of the six treatments. For thegraph titled “PT8 Up&Go” the y-axis is the time in seconds that it tookPT8 to stand from a seated position and walk a fixed distance that PT8walked each of the six times that the measurement was taken (eachparticipant tested also walked the same distance). As can be seen, PT8'stime to walk the distance decreased between the first measurement andthe last measurement indicating an improvement. For the graph titled“PT8 Balance” the y-axis is the time in seconds that PT8 was able tostand with one leg raised and where the maximum time measured was 120seconds. As can be seen, PT8's ability to balance on one leg increasedoverall through the course of the six treatments. For the graph titled“PT8 SF-12 Physical Health” the y-axis represents an SF-12 survey scorefor subjective physical health. As can be seen, PT8 showed an overallimprovement in scores over the course of the six treatments. For thegraph titled “PT8 SF-12 Emotional Health” the y-axis represents an SF-12survey score for subjective emotional health. As can be seen, PT8 showedan overall improvement in scores over the course of the six treatments.

FIG. 9 shows multiple graphs of macro data for a first sham participant(“SM1”) in the pilot study who is male and who received three shamplasmapheresis treatments (i.e. no plasmapheresis treatments weredelivered but participant was given illusion that plasmapheresis wasadministered). SM1 was the first of three participants who providedcontrol data that is presented here. SM1 had macro data collected beforeeach one of his three sham plasmapheresis treatments. The graphs of FIG.9 are titled to indicate which macro data they present. For each of thefive graphs shown in FIG. 9 , the x-axis has numbers 1 through 3indicating a time before each of the three sham plasmapheresistreatments when data was collected. For the graph titled “SM1 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, SM1'smeasured hand strength decreased overall over the course of the threesham treatments. For the graph titled “SM1 Up&Go” the y-axis is the timein seconds that it took SM1 to stand from a seated position and walk afixed distance that SM1 walked each of the three times that themeasurement was taken (each participant tested, including theplasmapheresis treated participants, also walked the same distance). Ascan be seen, SM1's time to walk the distance remained unchanged over theperiod of receiving sham plasmapheresis treatments. For the graph titled“SM1 Balance” the y-axis is the time in seconds that SM1 was able tostand with one leg raised and where the maximum time measured was 120seconds. As can be seen, SM1's ability to balance on one leg initiallydecreased and then increased to end overall unchanged through the courseof the three sham plasmapheresis treatments. For the graph titled “SM1SF-12 Physical Health” the y-axis represents an SF-12 survey score forsubjective physical health. As can be seen, SM1 showed an overallimprovement in scores over the course of the three sham plasmapheresistreatments. For the graph titled “SM1 SF-12 Emotional Health” the y-axisrepresents an SF-12 survey score for subjective emotional health. As canbe seen, SM1 showed an overall decrease in scores over the course of thethree sham plasmapheresis treatments.

FIG. 10 shows multiple graphs of macro data for a first sham participant(“SM2”) in the pilot study who is male and who received three shamplasmapheresis treatments (i.e. no plasmapheresis treatments weredelivered but participant was given illusion that plasmapheresis wasadministered). SM2 was the second of three participants who providedcontrol data that is presented here. SM2 had macro data collected beforeeach one of his three sham plasmapheresis treatments. The graphs of FIG.10 are titled to indicate which macro data they present. For each of thefive graphs shown in FIG. 10 , the x-axis has numbers 1 through 3indicating a time before each of the three sham plasmapheresistreatments when data was collected. For the graph titled “SM2 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, SM2'smeasured hand strength decreased overall over the course of the threesham treatments. For the graph titled “SM2 Up&Go” the y-axis is the timein seconds that it took SM2 to stand from a seated position and walk afixed distance that SM2 walked each of the three times that themeasurement was taken (each participant tested, including theplasmapheresis treated participants, also walked the same distance). Ascan be seen, SM2's time to walk the distance increased over the courseof the sham plasmapheresis treatments which indicates a worseningoverall. For the graph titled “SM2 Balance” the y-axis is the time inseconds that SM2 was able to stand with one leg raised and where themaximum time measured was 120 seconds. As can be seen, SM2 was able tostand the maximum measured time throughout course of receiving shamplasmapheresis treatments. For the graph titled “SM2 SF-12 PhysicalHealth” the y-axis represents an SF-12 survey score for subjectivephysical health. As can be seen, SM2 showed an overall improvement inscores over the course of the three sham plasmapheresis treatments. Forthe graph titled “SM1 SF-12 Emotional Health” the y-axis represents anSF-12 survey score for subjective emotional health. As can be seen, SM2showed an overall increase in scores over the course of the three shamplasmapheresis treatments.

FIG. 11 shows multiple graphs of macro data for a first sham participant(“SM3”) in the pilot study who is male and who received three shamplasmapheresis treatments (i.e. no plasmapheresis treatments weredelivered but participant was given illusion that plasmapheresis wasadministered). SM3 was the third of three participants who providedcontrol data that is presented here. SM3 had macro data collected beforeeach one of his three sham plasmapheresis treatments. The graphs of FIG.11 are titled to indicate which macro data they present. For each of thefive graphs shown in FIG. 11 , the x-axis has numbers 1 through 3indicating a time before each of the three sham plasmapheresistreatments when data was collected. For the graph titled “SM3 HandGrip”, the y-axis shows the measurements of grip strength in units of Kgmeasured with a standard grip strength device. As can be seen, SM3'smeasured hand strength decreased overall over the course of the threesham treatments. For the graph titled “SM3 Up&Go” the y-axis is the timein seconds that it took SM3 to stand from a seated position and walk afixed distance that SM3 walked each of the three times that themeasurement was taken (each participant tested, including theplasmapheresis treated participants, also walked the same distance). Ascan be seen, SM3's time to walk the distance was unchanged over thecourse of the sham plasmapheresis treatments. For the graph titled “SM3Balance” the y-axis is the time in seconds that SM3 was able to standwith one leg raised and where the maximum time measured was 120 seconds.As can be seen, SM3 had an overall decrease in time he was able tobalance on one leg over the course of the three sham plasmapheresistreatments. For the graph titled “SM3 SF-12 Physical Health” the y-axisrepresents an SF-12 survey score for subjective physical health. As canbe seen, SM3 showed an overall improvement in scores over the course ofthe three sham plasmapheresis treatments. For the graph titled “SM3SF-12 Emotional Health” the y-axis represents an SF-12 survey score forsubjective emotional health. As can be seen, SM3 showed an overalldecrease in scores over the course of the three sham plasmapheresistreatments.

In terms of timing of treatments, each of PT1, PT2, PT3, PT4, PT5, PT6,PT7, and PT8 received two treatments a month for three total months, andeach of the two monthly treatments for all eight participants werealways within at most one week of each other (i.e. at most one weekbetween treatment #1 and #2, treatment #3 and #4, and treatment #5 and#6). In addition, for PT3, there were 48 hours between treatment #3 andtreatment #4. For PT4, there were 48 hours between treatment #3 andtreatment #4. For PT5, there were 48 hours between treatment #1 andtreatment #2, there were 48 hours between treatment #3 and treatment #4,and there were 48 hours between treatment #5 and treatment #6. For PT6,there were 72 hours between treatment #3 and treatment #4, and therewere 48 hours between treatment #5 and treatment #6. For PT7, there were72 hours between treatment #1 and treatment #2, there were 48 hoursbetween treatment #3 and treatment #4, and there were 48 hours betweentreatment #5 and treatment #6. For PT5, there were 48 hours betweentreatment #1 and treatment #2, there were 48 hours between treatment #3and treatment #4, and there were 48 hours between treatment #5 andtreatment #6. As such, over half of the paired monthly treatmentsdelivered for PT1, PT2, PT3, PT4, PT5, PT6, PT7, and PT8 were within 72hours of one another.

TABLE III ID Hand Grip UP & Go Balance S-12/P SF-12/E PT1 6.5 −0.50(120) 15.68 21.75 PT2 2.5 −1.6 0(120) 5.91 0.98 PT3 −1.5 −2 20 4.389.33 PT4 14 −1.5 10(110)  3.3 6.52 PT5 6 −1 83 −1.54 22.17 PT6 0 −0.5 672.8 −0.93 PT7 −0.5 −2.5 0(120) −1.27 2.78 PT8 −3.5 −1 25 1.8 1.91 SM1−0.5 0 0(45)  6.98 −23.14 SM2 −1 0.5 0(120) 2.7 2.1 SM3 −3.5 0 −2 6.61−6.01

TABLE III above shows aggregate macro data for all 11 participants forwhich macro data is shown and described herein in FIGS. 1-11 and theirrespective accompanying descriptions. More specifically, the numericaldata in TABLE III is the difference (or delta) between the first datapoint and the last data point for each macro data parameter measured forall 11 participants. That is, for the eight participants that receivedsix plasmapheresis treatments each number in TABLE III is the lastmeasured data value (i.e. preceding the sixth plasmapheresis treatment)minus the first measured data value (i.e. preceding the firstplasmapheresis treatment). Similarly, for each of the three ham controlseach number is the last measured data value (i.e. preceding the thirdsham plasmapheresis treatment) minus the first measured data value (i.e.preceding the first sham plasmapheresis treatment). In addition, wherethere are numbers in parentheses, these numbers are the first measuredvalue. As can be seen for the participants, PT1, PT2, PT3, PT4, PT5,PT6, PT7, and PT8, there was across the board overall improvement in themultiple different metrics and especially with respect to the Up&Gometric where every treated patient showed improvement (for the Up&Go adecrease in value is an improvement). Compared to the three shamcontrols, SM1, SM2, and SM3, none of whom showed improvement instrength, walking, or balance, and far less overall improvement insubjective measures, there was strong indication that plasmapheresis hadpositive affect on strength, walking, balance, and mental wellbeingmetrics as compared to controls.

Micro Data

For the micro data, each study participant had three blood samples takenin the course of the plasmapheresis study, one before a firstplasmapheresis treatment, one in the middle of the plasmapheresistreatment cycle (i.e. before the fourth plasmapheresis treatment), andone before receiving the last plasmapheresis treatment. Blood sampleshad flow cytometry (including use of fluorescent coupled antibodies)performed on them to assess and quantify cell distribution as well ascell surface protein quantification (i.e. cell count of cells havingcell surface protein phenotype). Flow cytometry and analysis of bloodsamples of participants from the pilot study was performed at the BuckInstitute for Aging in Novato, California. For each sample, amounts ofexpression of the following cell surface markers (i.e. cell surfaceproteins) was measured: CD16, CD25, CD27, CD38, CD57, CD80, HLADR, IgM,KIR, KLRG1, NK1, NKg2a, and TIGIT. In addition, SA-β-gal, an indicatorof cellular senescence was quantified as well.

FIG. 12 shows graphs of normalized flow cytometry micro data assayingfor degree of expression of the cell surface markers CD16, CD25, CD27,CD38, CD57, CD80, HLADR, IgM, KIR, KLRG1, NK1, NKg2a, and TIGIT in cellsfrom blood samples taken from participants in the pilot study. An assaywas performed on expression of SA-β-gal as well in cells from bloodsamples taken from participants in the pilot study. The graphs in FIG.12 show micro data for blood samples taken from PT3. Three blood sampleswere taken from PT3, one before the first plasmapheresis treatment, onebefore the fourth plasmapheresis treatment, and one before the sixth andlast plasmapheresis treatment, and these are the x-values 1, 2, and 3 ofeach graph.

FIG. 13 shows graphs of normalized flow cytometry micro data assayingfor degree of expression of the cell surface markers CD16, CD25, CD27,CD38, CD57, CD80, HLADR, IgM, KIR, KLRG1, NK1, NKg2a, and TIGIT in cellsfrom blood samples taken from participants in the pilot study. An assaywas performed on expression of SA-β-gal as well in cells from bloodsamples taken from participants in the pilot study. The graphs in FIG.13 show micro data for blood samples taken from PT4. Three blood sampleswere taken from PT4, one before the first plasmapheresis treatment, onebefore the fourth plasmapheresis treatment, and one before the sixth andlast plasmapheresis treatment, and these are the x-values 1, 2, and 3 ofeach graph.

FIG. 14 shows graphs of normalized flow cytometry micro data assayingfor degree of expression of the cell surface markers CD16, CD25, CD27,CD38, CD57, CD80, HLADR, IgM, KIR, KLRG1, NK1, NKg2a, and TIGIT in cellsfrom blood samples taken from participants in the pilot study. An assaywas performed on expression of SA-β-gal as well in cells from bloodsamples taken from participants in the pilot study. The graphs in FIG.14 show micro data for blood samples taken from PT5. Three blood sampleswere taken from PT5, one before the first plasmapheresis treatment, onebefore the fourth plasmapheresis treatment, and one before the sixth andlast plasmapheresis treatment, and these are the x-values 1, 2, and 3 ofeach graph.

FIG. 15 shows graphs of normalized flow cytometry micro data assayingfor degree of expression of the cell surface markers CD16, CD25, CD27,CD38, CD57, CD80, HLADR, IgM, KIR, KLRG1, NK1, NKg2a, and TIGIT in cellsfrom blood samples taken from participants in the pilot study. An assaywas performed on expression of SA-β-gal as well in cells from bloodsamples taken from participants in the pilot study. The graphs in FIG.15 show micro data for blood samples taken from PT6. Three blood sampleswere taken from PT6, one before the first plasmapheresis treatment, onebefore the fourth plasmapheresis treatment, and one before the sixth andlast plasmapheresis treatment, and these are the x-values 1, 2, and 3 ofeach graph.

FIG. 16 shows graphs of normalized flow cytometry micro data assayingfor degree of expression of the cell surface markers CD16, CD25, CD27,CD38, CD57, CD80, HLADR, IgM, KIR, KLRG1, NK1, NKg2a, and TIGIT in cellsfrom blood samples taken from participants in the pilot study. An assaywas performed on expression of SA-β-gal as well in cells from bloodsamples taken from participants in the pilot study. The graphs in FIG.16 show micro data for blood samples taken from PT7. Three blood sampleswere taken from PT7, one before the first plasmapheresis treatment, onebefore the fourth plasmapheresis treatment, and one before the sixth andlast plasmapheresis treatment, and these are the x-values 1, 2, and 3 ofeach graph.

FIG. 17 shows graphs of normalized flow cytometry micro data assayingfor degree of expression of the cell surface markers CD16, CD25, CD27,CD38, CD57, CD80, HLADR, IgM, KIR, KLRG1, NK1, NKg2a, and TIGIT in cellsfrom blood samples taken from participants in the pilot study. An assaywas performed on expression of SA-β-gal as well in cells from bloodsamples taken from participants in the pilot study. The graphs in FIG.17 show micro data for blood samples taken from PT8. Three blood sampleswere taken from PT8, one before the first plasmapheresis treatment, onebefore the fourth plasmapheresis treatment, and one before the sixth andlast plasmapheresis treatment, and these are the x-values 1, 2, and 3 ofeach graph.

TABLE IV ID CD16 CD25 CD27 CD38 CD57 CD80 HLADR PT3 0.712408570.04503152 −0.0305988 0.00910674 −0.064439 −0.2246972 0.13545528 PT41.1129187 0.06767628 0.08079247 0.41053551 0.05358887 1.636585640.5941538 PT5 −0.2377705 −0.7618762 −0.2326512 −0.3667022 −0.21805971.16876294 −0.5084983 PT6 0.40331989 0.58625418 −0.2862214 −0.3193115−0.2878076 1.24095402 0.32989073 PT7 0.211982 −0.2566571 −0.1681396−0.2390395 −0.3137011 2.2009023 0.04759931 PT8 −0.2686451 −0.7306268−0.3970219 −0.3786687 −0.1898519 −0.4777714 −0.3679932 ID IgM* KIR KLRG1NK1 NKg2a* TIGIT SAbGal PT3 0.00069104 −0.2419516 −0.0754772 0.16079780.22066851 0.02742771 −0.0782061 PT4 1.34452284 0.97827381 −0.12184450.03809551 0.09519207 0.42050767 0.11329363 PT5 0.87019931 0.31031164−0.1434073 −0.1634698 −0.2987369 −0.4628126 −0.3486344 PT6 −0.70276361.28626122 −0.4544701 −0.2493025 0.29713981 −0.3410597 −0.1158705 PT71.32938848 0.47897551 −0.3687835 −0.1381616 −0.2044997 −0.0942918−0.3409115 PT8 −0.3768263 −0.4654095 −0.2407952 −0.2823279 −0.2643162−0.2939654 −0.4999459

TABLE IV above is broken into three parts to fit here and shows microdata for the six participants (PT3, PT4, PT5, PT6, PT7, and PT8) forwhich micro data is shown and described herein in FIGS. 12-17 and theirrespective accompanying descriptions. More specifically, the numericaldata in TABLE IV is the difference (or delta) between the first datapoint and the last data point for each micro data parameter measured forPT3, PT4, PT5, PT6, PT7, and PT8 who all received six plasmapheresistreatments. That is, for PT3, PT4, PT5, PT6, PT7, and PT8 each number inTABLE IV is the last measured data value (i.e. preceding the sixthplasmapheresis treatment) minus the first measured data value (i.e.preceding the first plasmapheresis treatment). In general, consensus forthe cell surface markers is that decreases in values indicate animprovement (i.e. through, at least, indicating a decrease ininflammation) except for the markers IgM and NKg2a which are marked withan asterisk to indicate that some research has indicated that anincrease in the values measured for these two markers indicatesimprovement. Values in TABLE IV that indicate improvement over thecourse of the plasmapheresis treatments delivered to PT3, PT4, PT5, PT6,PT7, and PT8 that indicate improvement are bolded in TABLE IV. As shown,PT5, PT6, PT7, and PT8 all had improvement in expression of cell surfacemarkers over the majority of markers measured between the first and lastblood samples. In addition looking at individual markers, KLRG1expression decreased in all participants shown in TABLE IV and CD27,CD57, and SAbGal expression decreased in almost all participants inTABLE IV.

General Statements Regarding this Disclosure

Each of the individual variations or embodiments described andillustrated herein has discrete components and features which may bereadily separated from or combined with the features of any of the othervariations or embodiments. Modifications may be made to adapt aparticular situation, material, composition of matter, process, processact(s) or step(s) to the objective(s), spirit or scope of the presentinvention.

Methods recited herein may be carried out in any order of the recitedevents that is logically possible, as well as the recited order ofevents. For example, the methods described herein do not necessarilyrequire the particular order of steps illustrated to achieve the desiredresult but rather one or more of the steps of a method as describedherein may be performed in a different order with respect to the othersteps. Moreover, additional steps or operations may be provided or stepsor operations may be eliminated to achieve the desired result.

It will be understood by one of ordinary skill in the art that all or aportion of the methods disclosed herein may be embodied in anon-transitory machine readable or accessible medium comprisinginstructions readable or executable by a processor or processing unit ofa computing device or other type of machine.

Furthermore, where a range of values is provided, every interveningvalue between the upper and lower limit of that range and any otherstated or intervening value in that stated range is encompassed withinthe invention. Also, any optional feature of the inventive variationsdescribed may be set forth and claimed independently, or in combinationwith any one or more of the features described herein.

All existing subject matter mentioned herein (e.g., publications,patents, patent applications and hardware) is incorporated by referenceherein in its entirety except insofar as the subject matter may conflictwith that of the present invention (in which case what is present hereinshall prevail). The referenced items are provided solely for theirdisclosure prior to the filing date of the present application.

Reference to a singular item includes the possibility that there areplural of the same items present. More specifically, as used herein andin the appended claims, the singular forms “a,” “an,” “said” and “the”include plural referents unless the context clearly dictates otherwise.It is further noted that the claims may be drafted to exclude anyoptional element. As such, this statement is intended to serve asantecedent basis for use of such exclusive terminology as “solely,”“only” and the like in connection with the recitation of claim elements,or use of a “negative” limitation. Unless defined otherwise, alltechnical and scientific terms used herein have the same meaning ascommonly understood by one of ordinary skill in the art to which thisinvention belongs.

This disclosure is not intended to be limited to the scope of theparticular forms set forth but is intended to cover alternatives,modifications, and equivalents of the variations or embodimentsdescribed herein. Further, the scope of the disclosure fully encompassesother variations or embodiments that may become obvious to those skilledin the art in view of this disclosure.

What is claimed is:
 1. A method of providing plasmapheresis to anindividual in order to improve a health status of the individual,comprising the steps of: (a) measuring, before an administration ofplasmapheresis, one or more of a strength of the individual, a balanceof the individual, a mental status of the individual, and a measure of awalking of the individual, thereby generating a pre-treatment healthstatus of the individual; (b) administering the plasmapheresis to theindividual; (c) measuring, following step (b), one or more of thestrength of the individual, the balance of the individual, the mentalstatus of the individual, and the measure of the walking of theindividual, thereby generating a post-treatment health status of theindividual; and (d) comparing the post-treatment health status of theindividual with the pre-treatment health status of the individual inorder to determine that the health status of the individual has improvedas a result of the plasmapheresis administration.
 2. The method of claim1, wherein step (a) occurs within 24 hours of the administering of theplasmapheresis to the individual in step (b).
 3. The method of claim 1,wherein the strength of the individual is measured in step (a), step(c), or steps (a) and (c) by measuring a grip strength of theindividual.
 4. The method of claim 1, wherein the balance of theindividual is measured in step (a), step (c), or steps (a) and (c) byhaving the individual stand on one leg and measuring how long theindividual remains standing with one leg raised.
 5. The method of claim1, wherein the mental status is measured in step (a), step (c), or steps(a) and (c) using a survey comprising questions that assess emotionalwellbeing.
 6. The method of claim 1, wherein the measure of the walkingof the individual is measured in step (a), step (c), or steps (a) and(c) by having the individual stand from a seated position and walk. 7.The method of claim 1, wherein the plasmapheresis that is administeredin step (b) exchanges at least one unit of plasma volume.
 8. The methodof claim 1, wherein the plasmapheresis that is administered in step (b)is administered over a plurality of treatment sessions.
 9. The method ofclaim 8, wherein two treatment sessions of the plurality of treatmentsessions are within 72 hours of each other.
 10. The method of claim 1,wherein the plasmapheresis that is administered in step (b) isadministered over a single treatment session.
 11. The method of claim 1,wherein the comparing the post-treatment health status of the individualwith the pre-treatment health status of the individual in step (d)results in a determination of a quantitative difference between thepost-treatment health status of the individual and the pre-treatmenthealth status of the individual.
 12. The method of claim 11, furthercomprising repeating steps (b)-(d) until the quantitative difference isa specific value.
 13. The method of claim 11, further comprisingrepeating steps (a)-(d) until the quantitative difference is a specificvalue.
 14. A method for treating a condition associated with aging in anindividual, comprising: (a) administering plasmapheresis to theindividual; and (b) monitoring for a change in the condition.
 15. Themethod of claim 14, wherein the condition associated with agingcomprises a loss of strength.
 16. The method of claim 15, wherein thestrength of the individual is monitored in step (b) by measuring a gripstrength of the individual.
 17. The method of claim 14, wherein thecondition associated with aging comprises a loss of balance.
 18. Themethod of claim 17, wherein balance of the individual is monitored instep (b) by having the individual stand on one leg and measuring howlong the individual remains standing with one leg raised.
 19. The methodof claim 14, wherein the condition associated with aging comprisesdiminished ability to walk.
 20. The method of claim 19, wherein theability to walk of the individual is monitored in step (b) by having theindividual stand from a seated position and walk.
 21. The method ofclaim 14, wherein the plasmapheresis that is administered in step (a)exchanges at least one unit of plasma volume.
 22. The method of claim14, wherein the plasmapheresis that is administered in step (a) isadministered over a plurality of treatment sessions.
 23. The method ofclaim 22, wherein two treatment sessions of the plurality of treatmentsessions are administered within 72 hours of each other.
 24. The methodof claim 14, wherein the plasmapheresis that is administered in step (a)is administered over a single treatment session.
 25. The method of claim14, further comprising repeating steps (a)-(b) until the change in thecondition is achieved.